Re: [ccp4bb] should the final model be refined against full datset

[Quyen Hoang]

Sorry, I don't quite understand your reasoning for how the structure  
is rendered useless if one refined it with all data.
Would your argument also apply to all the structures that were refined  
before R-free existed?


Quyen



You should enter the statistics for the model and data that you  
actually deposit, not statistics for some other model that you might  
have had at one point but which the PDB will never see.  Not only  
does refining against R-free make it impossible to verify and  
validate your structure, it also means that any time you or anyone  
else wants to solve an isomorphous structure by MR using your  
structure as a search model, or continue the refinement with higher- 
resolution data, you will be starting with a model that has been  
refined against all reflections.  So any future refinements done  
with that model against isomorphous data are pre-biased, making your  
model potentially useless.


I'm amazed that anyone is still depositing structures refined  
against all data, but the PDB does still get a few.  The benefit of  
including those extra 5% of data is always minimal in every paper  
I've seen that reports such a procedure, and far outweighed by  
having a reliable and relatively unbiased validation statistic that  
is preserved in the final deposition.  (The situation may be  
different for very low resolution data, but those structures are a  
tiny fraction of the PDB.)


-Nat

Re: [ccp4bb] should the final model be refined against full datset

[Quyen Hoang]

I still don't understand how a structure model refined with all data  
would negatively affect the determination and/or refinement of an  
isomorphous structure using a different data set (even without doing  
SA first).


Quyen

On Oct 14, 2011, at 4:35 PM, Nat Echols wrote:

On Fri, Oct 14, 2011 at 1:20 PM, Quyen Hoang   
wrote:
Sorry, I don't quite understand your reasoning for how the structure  
is rendered useless if one refined it with all data.


"Useless" was too strong a word (it's Friday, sorry).  I guess  
simulated annealing can address the model-bias issue, but I'm not  
totally convinced that this solves the problem.  And not every  
crystallographer will run SA every time he/she solves an isomorphous  
structure, so there's a real danger of misleading future users of  
the PDB file.  The reported R-free, of course, is still meaningless  
in the context of the deposited model.


Would your argument also apply to all the structures that were  
refined before R-free existed?


Technically, yes - but how many proteins are there whose only  
representatives in the PDB were refined this way?  I suspect very  
few; in most cases, a more recent model should be available.


-Nat

Re: [ccp4bb] should the final model be refined against full datset

[Quyen Hoang]

Thanks for the clear explanation. I understood that.
But I was trying to understand how this would negatively affects the  
initial model to render it useless or less useful.
In the scenario that you presented, I would expect a better result  
(better model) if the initial model was refined with all data, thus  
more useful.
Sure, again in your scenario, the "new" structure has seen R-free  
reflections in the equivalent indexes of its replacement model, but  
their intensities should be different anyway, so I am not sure how  
this is bad. Even if the bias is huge, let's say this bias results in  
1% reduction in initial R-free (exaggerating here), how would this  
makes one's model bad or how would this be bad for one's science?
In the end, our objective is to build the best model possible and I  
think that more data would likely result in better model, not the  
other way around. If we can agree that refining a model with all data  
would result in a better model, then wouldn't not doing so constitute  
a compromise of model quality for a more "pure" statistic?


I had not refined a model with all data before (just to keep inline),  
but I  wondered if I was doing the best thing.


Cheers,
Quyen
On Oct 14, 2011, at 5:27 PM, Phil Jeffrey wrote:

Let's say you have two isomorphous crystals of two different protein- 
ligand complexes.  Same protein different ligand, same xtal form.   
Conventionally you'd keep the same free set reflections (hkl values)  
between the two datasets to reduce biasing.  However if the first  
model had been refined against all reflections there is no longer a  
free set for that model, thus all hkl's have seen the atoms during  
refinement, and so your R-free in the second complex is initially  
biased to the model from the first complex. [*]


The tendency is to do less refinement in these sort of isomorphous  
cases than in molecular replacement solutions, because the  
structural changes are usually far less (it is isomorphous after  
all) so there's a risk that the R-free will not be allowed to fully  
float free of that initial bias.  That makes your R-free look better  
than it actually is.


This is rather strongly analogous to using different free sets in  
the two datasets.


However I'm not sure that this is as big of a deal as it is being  
made to sound.  It can be dealt with straightforwardly.  However  
refining against all the data weakens the use of R-free as a  
validation tool for that particular model so the people that like to  
judge structures based on a single number (i.e. R-free) are going to  
be quite put out.


It's also the case that the best model probably *is* the one based  
on a careful last round of refinement against all data, as long as  
nothing much changes.  That would need to be quantified in some  
way(s).


Phil Jeffrey
Princeton

[* Your R-free is also initially model-biased in cases where the  
data are significant non-isomorphous or you're using two different  
xtal forms, to varying extents]





I still don't understand how a structure model refined with all data
would negatively affect the determination and/or refinement of an
isomorphous structure using a different data set (even without  
doing SA

first).

Quyen

On Oct 14, 2011, at 4:35 PM, Nat Echols wrote:


On Fri, Oct 14, 2011 at 1:20 PM, Quyen Hoang mailto:qqho...@gmail.com>> wrote:

   Sorry, I don't quite understand your reasoning for how the
   structure is rendered useless if one refined it with all data.


"Useless" was too strong a word (it's Friday, sorry). I guess
simulated annealing can address the model-bias issue, but I'm not
totally convinced that this solves the problem. And not every
crystallographer will run SA every time he/she solves an isomorphous
structure, so there's a real danger of misleading future users of  
the
PDB file. The reported R-free, of course, is still meaningless in  
the

context of the deposited model.

   Would your argument also apply to all the structures that were
   refined before R-free existed?


Technically, yes - but how many proteins are there whose only
representatives in the PDB were refined this way? I suspect very  
few;

in most cases, a more recent model should be available.

-Nat

Re: [ccp4bb] what to do with disordered side chains

[Quyen Hoang]

I don't have strong preference either way, but if one has good density  
for backbone and no density for a corresponding side-chain, would it  
be reasonable to believe that the side-chain might actually exists and  
that it has no visible density at a certain contour level might be  
because it was moving around (or static disorder)? And if B-factor is  
an estimate of thermo-motion (or static disorder), then would it not  
be reasonable to accept that building the side-chain and let B-factor  
sky rocket might reflect reality more so than not building it?


Cheers,
Quyen
___
Quyen Hoang, Ph.D
Assistant Professor
Department of Biochemistry and Molecular Biology,
Stark Neurosciences Research Institute
Indiana University School of Medicine
635 Barnhill Drive, Room MS0013D
Indianapolis, Indiana 46202-5122

Phone: 317-274-4371
Fax: 317-274-4686
email: qqho...@iupui.edu
Website: www.hoanglab.com



On Mar 30, 2011, at 2:04 PM, James Holton wrote:



I'm afraid this is not a problem that can be solved by  
"standardization".


Fundamentally, if you are a scientist who has collected some data  
(be it diffraction spot intensities, cell counts, or substrate  
concentration vs time), and you have built a "model" to explain that  
data (be it a constellation of atoms in a unit cell, exponential  
population growth, or a microscopic reaction mechanism), I think it  
is generally expected that your model explain the data "to within  
experimental error".  Unfortunately, this is never the case in  
macromolecular crystallography, where the model-data disagreement  
(Fobs-Fcalc) is ~4-5x bigger than the "error bars" (sigma(F)).


Now, there is nothing shameful about an incomplete model, especially  
when thousands of very intelligent people working over half a  
century have not been able to come up with a better way to build  
one.  In fact, perhaps a better name for the "disordered side chain  
problem" would be "dark density"?  This name would place it properly  
amongst "dark matter", "dark energy" and other fudge factors  
introduced to try and explain why our "standard model" is not  
consistent with observation?  That is, "dark density" is the stuff  
we can't see, but nonetheless must be there somewhere.


Whatever it is, I personally do hold a vain belief that perhaps  
someday soon the problem of "dark density" will be solved, and that  
presently instituting a "policy" requiring that all macromolecular  
models from this day forward remain at least as incomplete as  
yesterday's models is not a very good idea.  I say: if you think  
there is "something there" then you should build it in, especially  
if it is important to the conclusions you are trying to make.  You  
can defend your model the same way you would defend any other  
scientific model: by using established statistics to show that it  
agrees with the data better than an "alternative model" (like  
leaving it out).  It is YOUR model, after all!  Only you are  
responsible for how "right" it is.


I do appreciate that students and other novices may have a harder  
time defining "surfaces" and measuring hydrogen bond lengths in  
these pesky "floppy regions", but perhaps their education would be  
served better by learning the truth sooner than later?


-James Holton
MAD Scientist


On 3/30/2011 9:26 AM, Filip Van Petegem wrote:


Hello Mark,

I absolutely agree with this.  The worst thing is when everybody is  
following their own personal rules, and there are no major  
guidelines for end-users to figure out how to interpret those  
parts.  I assume there are no absolute guidelines simply because  
there isn't any consensus among crystallographers... (from what we  
can gather from this set of emails...). On the other hand, this  
discussion has flared up many times in the past, and maybe it's  
time for a powerful dictator at the PDB to create the law...


Filip Van Petegem



On Wed, Mar 30, 2011 at 8:37 AM, Mark J van Raaij > wrote:
perhaps the IUCr and/or PDB (Gerard K?) should issue some  
guidelines along these lines?

And oblige us all to follow them?
Mark J van Raaij
Laboratorio M-4
Dpto de Estructura de Macromoleculas
Centro Nacional de Biotecnologia - CSIC
c/Darwin 3, Campus Cantoblanco
E-28049 Madrid, Spain
tel. (+34) 91 585 4616
http://www.cnb.csic.es/content/research/macromolecular/mvraaij/index.php?l=1



On 30 Mar 2011, at 17:29, Phoebe Rice wrote:

> I've now polled 4 fairly savvy "end users" of crystal structures  
and there seems to be a consensus:

>
> - they all know what B is and how to look for regions of high B  
(with, say, pymol) and they know not to make firm conclusions about  
H-bonds to flaming red side chains.
> - None of them would ever think to look at occup

Re: [ccp4bb] what to do with disordered side chains

[Quyen Hoang]

We are getting off topic a little bit.

Original topic: is it better to not build disordered sidechains or  
build them and let B-factors take care of it?

Ed's poll got almost a 50:50 split.
Question still unanswered.

Second topic introduced by Pavel: "Your B-factors are valid within a  
harmonic (small) approximation of atomic vibrations. Larger scale  
motions you are talking about go beyond the harmonic approximation,  
and using the B-factor to model them is abusing the corresponding  
mathematical model."
And that these large scale motions (disorders) are better represented  
by "alternative conformations and associated with them occupancies".


My question is, how many people here do this?
If you're currently doing what Pavel suggested here, how do you decide  
where to keep the upper limit of B-factors and what the occupancies  
are for each atom (data with resolution of 2.0A or worse)? I mean, do  
you cap the B-factor at a reasonable number to represent natural  
atomic vibrations (which is very small as Pavel pointed out) and then  
let the occupancies pick up the slack? More importantly, what is your  
reason for doing this?


Cheers and thanks for your contribution,
Quyen


On Mar 30, 2011, at 5:20 PM, Pavel Afonine wrote:


Mark,
alternative conformations and associated with them occupancies are  
to describe the larger scale disorder (the one that goes beyond the  
B-factor's capability to cope with).

Multi-model PDB files is another option.
Best,
Pavel.


On Wed, Mar 30, 2011 at 2:15 PM, VAN RAAIJ , MARK JOHAN > wrote:
yet, apart from (and additionally to) modelling two conformations of  
the side-chain, the B-factor is the only tool we have (now).


Quoting Pavel Afonine:

> Hi  Quyen,
>
>
> (...) And if B-factor is an estimate of thermo-motion (or static  
disorder),
>> then would it not be reasonable to accept that building the side- 
chain and
>> let B-factor sky rocket might reflect reality more so than not  
building it?

>>
>
> NO.  Your B-factors are valid within a harmonic (small)  
approximation of
> atomic vibrations. Larger scale motions you are talking about go  
beyond the
> harmonic approximation, and using the B-factor to model them is  
abusing the

> corresponding mathematical model.
> http://www.phenix-online.org/newsletter/CCN_2010_07.pdf
>
> Pavel.
>

Mark J van Raaij
Laboratorio M-4
Dpto de Estructura de Macromoléculas
Centro Nacional de Biotecnología - CSIC

c/Darwin 3, Campus Cantoblanco
28049 Madrid
tel. 91 585 4616
email: mjvanra...@cnb.csic.es

Re: [ccp4bb] what to do with disordered side chains

[Quyen Hoang]

Thank you for your post, Herman.
Since there is no holy bible to provide guidance, perhaps we should  
hold off the idea of electing a "powerful dictator" to enforce this?
- at least until we all can come to a consensus on how the "dictator"  
should dictate...


Cheers,
Quyen


On Mar 31, 2011, at 10:22 AM, herman.schreu...@sanofi-aventis.com wrote:


Dear Quyen,
I am afraid you won't get any better answers than you got so far.  
There is no holy bible telling you what to do with disordered side  
chains. I fully agree with James that you should try to get the best  
possible model, which best explains your data and that will be your  
decision. Here are my 2 cents:


-If you see alternative positions, you have to build them.
-If you do not see alternative positions, I would not replace one  
fantasy (some call it most likely) orientation with 2 or 3 fantasy  
orientations.
-I personally belong to the "let the B-factors take care of it"  
camp, but that is my personal opinion. Leaving side chains out could  
lead to misinterpretations by slightly less savy users of our data,  
especially when charge distributions are being studied. Besides, we  
know (almost) for sure that the side chain is there, it is only  
disordered and as we just learned, even slightly less savy users  
know what flaming red side chains mean. Even if they may not be  
mathematically entirely correct, huge B-factors clearly indicate  
that there is disorder involved.
-I would not let occupancies take up the slack since even very savy  
users have never heard of them and again, the side chain is fully  
occupied, only disordered. Of course if you build alternate  
positions, you have to divede the occupancies amongst them.


Best,
Herman

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf  
Of Quyen Hoang

Sent: Thursday, March 31, 2011 3:55 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] what to do with disordered side chains

We are getting off topic a little bit.

Original topic: is it better to not build disordered sidechains or  
build them and let B-factors take care of it?

Ed's poll got almost a 50:50 split.
Question still unanswered.

Second topic introduced by Pavel: "Your B-factors are valid within a  
harmonic (small) approximation of atomic vibrations. Larger scale  
motions you are talking about go beyond the harmonic approximation,  
and using the B-factor to model them is abusing the corresponding  
mathematical model."
And that these large scale motions (disorders) are better  
represented by "alternative conformations and associated with them  
occupancies".


My question is, how many people here do this?
If you're currently doing what Pavel suggested here, how do you  
decide where to keep the upper limit of B-factors and what the  
occupancies are for each atom (data with resolution of 2.0A or  
worse)? I mean, do you cap the B-factor at a reasonable number to  
represent natural atomic vibrations (which is very small as Pavel  
pointed out) and then let the occupancies pick up the slack? More  
importantly, what is your reason for doing this?


Cheers and thanks for your contribution,
Quyen


On Mar 30, 2011, at 5:20 PM, Pavel Afonine wrote:


Mark,
alternative conformations and associated with them occupancies are  
to describe the larger scale disorder (the one that goes beyond the  
B-factor's capability to cope with).

Multi-model PDB files is another option.
Best,
Pavel.


On Wed, Mar 30, 2011 at 2:15 PM, VAN RAAIJ , MARK JOHAN > wrote:
yet, apart from (and additionally to) modelling two conformations  
of the side-chain, the B-factor is the only tool we have (now).


Quoting Pavel Afonine:

> Hi  Quyen,
>
>
> (...) And if B-factor is an estimate of thermo-motion (or static  
disorder),
>> then would it not be reasonable to accept that building the side- 
chain and
>> let B-factor sky rocket might reflect reality more so than not  
building it?

>>
>
> NO.  Your B-factors are valid within a harmonic (small)  
approximation of
> atomic vibrations. Larger scale motions you are talking about go  
beyond the
> harmonic approximation, and using the B-factor to model them is  
abusing the

> corresponding mathematical model.
> http://www.phenix-online.org/newsletter/CCN_2010_07.pdf
>
> Pavel.
>

Mark J van Raaij
Laboratorio M-4
Dpto de Estructura de Macromoléculas
Centro Nacional de Biotecnología - CSIC

c/Darwin 3, Campus Cantoblanco
28049 Madrid
tel. 91 585 4616
email: mjvanra...@cnb.csic.es

Re: [ccp4bb] what to do with disordered side chains

[Quyen Hoang]

Dear Gerard,

I agree with you based on debates at some conferences.

But, based on what I have seen here so far, it seems to me that everybody knows 
exactly what to do with disordered side chains.
People that want to build structures to best fit the data tend to prefer 
omitting disordered side chains. On the other hand, people that want to build 
structures to best represent reality tend to prefer building them. I don't see 
any disagreement here nor do I see any problems with either approach. Different 
people collect the same data to study different things and I feel that they are 
entitle to view and interpret the data the way that they fine most meaningful. 

Equations are attempts to describe reality, I don't see why we should constrain 
reality to fit equations. 

Cheers,
Quyen


On Mar 31, 2011, at 12:21 PM, Gerard Bricogne wrote:

> Dear Quyen,
> 
> On Thu, Mar 31, 2011 at 11:27:58AM -0400, Quyen Hoang wrote:
>> Thank you for your post, Herman.
>> Since there is no holy bible to provide guidance, perhaps we should hold 
>> off the idea of electing a "powerful dictator" to enforce this?
>> - at least until we all can come to a consensus on how the "dictator" 
>> should dictate...
>> 
> 
> ... but that might well be even harder than to decide what to do with
> disordered side chains ... .
> 
> 
> With best wishes,
> 
>  Gerard.
> 
> --
> 
> ===
> * *
> * Gerard Bricogne g...@globalphasing.com  *
> * *
> * Global Phasing Ltd. *
> * Sheraton House, Castle Park Tel: +44-(0)1223-353033 *
> * Cambridge CB3 0AX, UK   Fax: +44-(0)1223-366889 *
> * *
> ===
>> 
>> 
>> On Mar 31, 2011, at 10:22 AM, herman.schreu...@sanofi-aventis.com wrote:
>> 
>>> Dear Quyen,
>>> I am afraid you won't get any better answers than you got so far. There is 
>>> no holy bible telling you what to do with disordered side chains. I fully 
>>> agree with James that you should try to get the best possible model, which 
>>> best explains your data and that will be your decision. Here are my 2 
>>> cents:
>>> 
>>> -If you see alternative positions, you have to build them.
>>> -If you do not see alternative positions, I would not replace one fantasy 
>>> (some call it most likely) orientation with 2 or 3 fantasy orientations.
>>> -I personally belong to the "let the B-factors take care of it" camp, but 
>>> that is my personal opinion. Leaving side chains out could lead to 
>>> misinterpretations by slightly less savy users of our data, especially 
>>> when charge distributions are being studied. Besides, we know (almost) for 
>>> sure that the side chain is there, it is only disordered and as we just 
>>> learned, even slightly less savy users know what flaming red side chains 
>>> mean. Even if they may not be mathematically entirely correct, huge 
>>> B-factors clearly indicate that there is disorder involved.
>>> -I would not let occupancies take up the slack since even very savy users 
>>> have never heard of them and again, the side chain is fully occupied, only 
>>> disordered. Of course if you build alternate positions, you have to divede 
>>> the occupancies amongst them.
>>> 
>>> Best,
>>> Herman
>>> 
>>> From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of 
>>> Quyen Hoang
>>> Sent: Thursday, March 31, 2011 3:55 PM
>>> To: CCP4BB@JISCMAIL.AC.UK
>>> Subject: Re: [ccp4bb] what to do with disordered side chains
>>> 
>>> We are getting off topic a little bit.
>>> 
>>> Original topic: is it better to not build disordered sidechains or build 
>>> them and let B-factors take care of it?
>>> Ed's poll got almost a 50:50 split.
>>> Question still unanswered.
>>> 
>>> Second topic introduced by Pavel: "Your B-factors are valid within a 
>>> harmonic (small) approximation of atomic vibrations. Larger scale motions 
>>> you are talking about go beyond the harmonic approximation, and using the 
>>> B-factor to model them is abusing the corresponding mathematical model."
>>> And that these large scale motions (disorders) a

Re: [ccp4bb] Crystallographic Breakthrough - DarkMatter Version 1.0

[Quyen Hoang]

Will this affect my reprocessing of the data with D*TREK on my journey  
to XPLORE the planets MERCURY and rPLUTO in my ENDEAVOUR to find and  
BUSTER some CRYSTALS with my on-board TNT into XPOWDER?
I am still trying to GRASP the idea of AUTODOCKing on precise HKL  
locations based on the SHARP but CONVX images produced by CRYSTAL  
STUDIO.


Quyen


On Apr 1, 2011, at 2:06 AM, Ethan Merritt wrote:


Hi to all on ccp4bb:

What better day to announce the availability of a breakthrough  
technique

in macromolecular crystallography?

Given recent discussion and in particular James Holton's suggestion  
that
the problem of disordered sidechains is a problem akin to the  
difficulty

of describing dark matter and dark energy...

I am happy to announce a new crystallographic tool that can improve  
your
model by accounting for an often-neglected physical property. A  
detailed
explanation, references, and a preliminary implementation of the  
program

can be downloaded from

http://skuld.bmsc.washington.edu/DarkMatter

--
Ethan A Merritt
Karmic Diffraction Project
Fine crystallography since April 1, 2011
"What goes around, comes around - usually as a symmetry equivalent"

Re: [ccp4bb] what to do with disordered side chains

[Quyen Hoang]

If these users exist (I don't doubt that they do), then they would also might 
think that lysine residues sometimes look identical to alanine - if the atoms 
after beta carbon of the lysine are deleted in the PDB due to lack of density.

So, I guess, if one's objectives in solving structures are to provide these 
users with coordinates that they could us, then it would make more sense to me 
to find out what one's "customers" want, rather than speculating about it. Or 
at least, train one's "customers" how to use one's products - I believe that's 
what people in business do.

However, I think that many people solve structures for their own consumption - 
they are their own customers - therefore, it's really up to them to cook it 
anyway they find most tasteful. Others can agree or disagree, but we know that 
not everybody has the same taste.

Cheers,
Quyen


On Apr 3, 2011, at 12:54 AM, Prof. Joel L. Sussman wrote:

> I think Frances is right, i.e. most non crystallographers ignore 
> "anything beyond the x, y, z coordinates (i.e. beyond column 54)"
> [as Frances wrote]. 
> Thus if a crystallographer put in coords that he/she does NOT see,
> even with OCC=0, or an enormously large Bfactor, these coords are usually 
> treated in just the same way that experimentally observed coords are treated. 
> Thus my feeling is that if one does NOT see the coords in the electron
> density, they should NOT be included, and let someone else try to model 
> them in, but they should be aware that they are modeling them.
> Joel L. Sussman
> 
> 
> 
> On 3 Apr 2011, at 06:15, Frances C. Bernstein wrote:
> 
>> Doing something sensible in the major software packages, both
>> for graphics and for other analysis of the structure, could
>> solve the problem for most users.
>> 
>> But nobody knows what other software is out there being used by
>> individuals or small groups.  And the more remote the authors
>> of that software are from protein structure solution the more
>> likely it is that they have not/will not properly handle atoms
>> with zero occupancy or high B values, for example.
>> 
>> I am absolutely positive that there is software that does its
>> voodoo on ATOM/HETATM records and pays absolutely no attention
>> to anything beyond the x, y, z coordinates (i.e. beyond column 54).
>> 
>>Frances Bernstein
>> 
>> =
>> Bernstein + Sons
>> *   *   Information Systems Consultants
>> 5 Brewster Lane, Bellport, NY 11713-2803
>> *   * ***
>>  *Frances C. Bernstein
>>  *   ***  f...@bernstein-plus-sons.com
>> *** *
>>  *   *** 1-631-286-1339FAX: 1-631-286-1999
>> =
>> 
>> On Sat, 2 Apr 2011, Jacob Keller wrote:
>> 
 I guess I missed it in the flurry of replies to this thread over the
 last few days, but what exactly is so terrible about keeping the atoms
 (since you have chemical evidence from protein sequence that they are
 there, and even if there is X-ray damage they were originally there and
 are likely still there in a subset of the molecules), but changing
 occupancy to zero as an acknowledgment that your data does not provide
 evidence to support a specific atomic position for these atoms?
>>> 
>>> Some users might pull up the structure, see those atoms, and think
>>> their positions were based on data, which they were not, and then draw
>>> conclusions based on them. I agree that occ=0 is tantamount to the
>>> suggestion you queried, however.
>>> 
>>> A somewhat key question might be: across the various molecular
>>> visualization programs, what is the default way to handle atoms with
>>> occ=0? Perhaps those programs might be the best place to fix the
>>> problem...
>>> 
>>> JPK
>>> 
>>> 
>>> ***
>>> Jacob Pearson Keller
>>> Northwestern University
>>> Medical Scientist Training Program
>>> cel: 773.608.9185
>>> email: j-kell...@northwestern.edu
>>> ***
>>> 
> 

Re: [ccp4bb] Crystallographic Breakthrough - DarkMatter Version 1.0

[Quyen Hoang]

This could be my last post ...

I am now leaving mercury
It's surface was safe and free
Meet a mystical gryphon and some alien mosquitos
They gave me some protein crystals
Which tried to shoot like 007
Missed them all but the heaven
Will try again with my new X8
Which 007 would surely hate
Now approaching pluto
Visibility is really low
My landing site had became flatter
And the uncertainty perimeter around it had turned into darkmatter
If I miss the apex of my landing site
Will I disappear right out of sight
Will I turn into an infinite spaghetti
Which I am sure the novice users would get all gitty

Cheers,
Quyen


On Apr 4, 2011, at 5:19 AM, Eleanor Dodson wrote:


Very clever..
Eleanor


On 04/01/2011 03:19 PM, Quyen Hoang wrote:
Will this affect my reprocessing of the data with D*TREK on my  
journey

to XPLORE the planets MERCURY and rPLUTO in my ENDEAVOUR to find and
BUSTER some CRYSTALS with my on-board TNT into XPOWDER?
I am still trying to GRASP the idea of AUTODOCKing on precise HKL
locations based on the SHARP but CONVX images produced by CRYSTAL  
STUDIO.


Quyen


On Apr 1, 2011, at 2:06 AM, Ethan Merritt wrote:


Hi to all on ccp4bb:

What better day to announce the availability of a breakthrough  
technique

in macromolecular crystallography?

Given recent discussion and in particular James Holton's  
suggestion that
the problem of disordered sidechains is a problem akin to the  
difficulty

of describing dark matter and dark energy...

I am happy to announce a new crystallographic tool that can  
improve your
model by accounting for an often-neglected physical property. A  
detailed
explanation, references, and a preliminary implementation of the  
program

can be downloaded from

http://skuld.bmsc.washington.edu/DarkMatter

--
Ethan A Merritt
Karmic Diffraction Project
Fine crystallography since April 1, 2011
"What goes around, comes around - usually as a symmetry equivalent"

Re: [ccp4bb] Nanodrop versus Nanophotomter Pearl versus good old Bradford.

[Quyen Hoang]

We also have not experienced any problems with a Nanodrop 2000C.
No one in my touched the two boxes of Bradford and BCA kits that we  
have, because we have been very happy with the Nanodrop.


Quyen
___
Quyen Hoang, Ph.D
Assistant Professor
Department of Biochemistry and Molecular Biology,
Stark Neurosciences Research Institute
Indiana University School of Medicine
635 Barnhill Drive, Room MS0013D
Indianapolis, Indiana 46202-5122

Phone: 317-274-4371
Fax: 317-274-4686
email: qqho...@iupui.edu
Website: www.hoanglab.com



On Jun 16, 2011, at 3:54 PM, Francis E Reyes wrote:

Never had problems with evaporation (and this is in the relatively  
dry climate of Denver, CO, especially in the winter when the  
relative humidity is in the low 20%).


Using the Thermo Scientific Nanodrop 2000c.

We use it also as a prerequisite for ITC, which can be very  
sensitive to proper concentrations.


F


On Jun 16, 2011, at 1:15 PM, Arnon Lavie wrote:

Dear fellow crystallographers - a question about spectrophotometers  
for protein concentration determination.


We are so last millennium - using Bradford reagent/ 1 ml cuvette  
for protein conc. determination.


We have been considering buying a Nanodrop machine (small volume,  
no dilution needed, fast, easy).
However, while testing our samples using a colleague's machine, we  
have gotten readings up to 100% different to our Bradford assay  
(all fully purified proteins). For example, Bradford says 6 mg/ml,  
Nanodrop 3 mg/ml. So while it is fun/easy to use the Nanodrop, I am  
not sure how reliable are the measurements (your thoughts?).


So QUESTION 1: What are people's experience regarding the  
correlation between Nanodrop and Bradford?


While researching the Nanodrop machine, I heard about the Implen  
NanoPhotmeter Pearl.
So Question 2: Is the Pearl better/worse/same as the Nanodrop for  
our purpose?


Thank you for helping us to advance to the next millennium, even if  
it is nearly a dozen years late.


Arnon

--
***
Arnon Lavie, Professor
Dept. of Biochemistry and Molecular Genetics
University of Illinois at Chicago
900 S. Ashland Ave.
Molecular Biology Research Building, Room 1108 (M/C 669)
Chicago, IL 60607
U.S.A.
   Tel:(312) 355-5029
   Fax:(312) 355-4535
   E-mail: la...@uic.edu
   http://www.uic.edu/labs/lavie/
***

[ccp4bb] Postdoc Position in Molecular Neuroscience

[Quyen Hoang]

A postdoctoral position is available to study the mechanisms of  
Parkinson disease - please see attached file for detail.


Genetic studies in the past decade have identified a number of genes  
as causal factors of Parkinson disease. Our research focuses on  
understanding the biochemical functions of the disease associated  
proteins and the biological pathways in which they function. Our core  
research technique is structural biology (protein X-ray  
crystallography) and structure-based drug design, but we use a wide  
range of techniques including, computational chemistry, biophysical  
techniques, enzymology, molecular biology and cell biology.


 Our laboratory is affiliated with the Department of Biochemistry and  
Molecular Biology (http://www.medicine.iu.edu/body.cfm?id=998) as well  
as the Stark Neurosciences Research Institute (http:// 
snri.iusm.iu.edu/) at IUPUI (http://www.iupui.edu/) in Indianapolis  
Indiana (http://en.wikipedia.org/wiki/Indianapolis).


 We are looking for an enthusiastic person who holds a PhD in  
biochemistry or related field, has a strong background in molecular  
biology and protein biochemistry. An ideal candidate should have  
genuine interest in science and desire to learn new techniques to  
tackle challenging and important questions.


 Please email a cover letter, CV, and three reference letters to:

qqho...@iupui.edu

Thank you for looking and happy holidays!

Cheers,
Quyen
___
Quyen Hoang, Ph.D
Department of Biochemistry and Molecular Biology,
Stark Neurosciences Research Institute
Indiana University School of Medicine
635 Barnhill Drive, Room MS0013D
Indianapolis, Indiana 46202-5122

Phone: 317-274-4371
Fax: 317-274-4686
email: qqho...@iupui.edu

Re: [ccp4bb] Reg. Protein purification

[Quyen Hoang]

I have seen this when expressing sub-domains of a larger protein.
Adding some (about 0.5% w/v) BOG (beta octylglucoside) worked well for  
cases similar to what you've described.


Cheers,
Quyen

___
Quyen Hoang, Ph.D
Department of Biochemistry and Molecular Biology,
Stark Neurosciences Research Institute
Indiana University School of Medicine
635 Barnhill Drive, Room MS0013D
Indianapolis, Indiana 46202-5122

Phone: 317-274-4371
Fax: 317-274-4686
email: qqho...@iupui.edu

On Jan 8, 2010, at 7:41 AM, Sivaraman Padavattan wrote:


Dear all,
I am trying to express the human protein using bacterial expression  
strain (Rosetta) and purified using Ni-NTA affinity purification.  
The Molecular weight of out protein is 47 kDa. In SDS-PAGE, we have  
seen that 27 kDa contaminant protein co-eluted with our protein even  
at high concentration of Imidazole. In Superose 12 column, these two  
proteins eluted as a single peak and its corresponding molecular  
weight suggestive of partial interaction. By mass mapping we have  
found 27 kDa band is an adenylate kinase. Is there any specific way  
to separate adenylate kinase  from our protein?

Thanks in advance,

Sivaraman Padavattan

Re: [ccp4bb] Deposition of riding H: R-factor is overrated

[Quyen Hoang]

As a relatively inexperienced scientist, I find this discussion  
fascinating.
I wonder if NMR and EM people are also worried about depositing enough  
"modeled info" to allow back calculation of data.


Regarding the original discussion of whether to deposit riding  
hydrogens used in the refinement, it seems that we are trying to  
deposit one model to satisfy two different purposes - one for model  
validation and the other for model interpretation (use in docking  
etc), and what's good for one purpose might not be necessarily good  
for the other.
I wonder if it would help to deposit two different models; one  
precisely reflects the model used in refinement and the other an  
energy minimized model with predicted hydrogens and alternative  
conformations removed?


Cheers,
Quyen

______
Quyen Hoang, Ph.D
Assistant Professor
Department of Biochemistry and Molecular Biology,
Stark Neurosciences Research Institute
Indiana University School of Medicine
635 Barnhill Drive, Room MS0013D
Indianapolis, Indiana 46202-5122

Phone: 317-274-4371
Fax: 317-274-4686
email: qqho...@iupui.edu


On Sep 17, 2010, at 8:28 AM, Ian Tickle wrote:


Oh, goodness, I see: even here, we would need clear rules what the
calculated structure factors are, which weights are were, which  
bulk solvent

correction was applied ... a maze, too!


Fortunately the X-ray & restraint weights/target values are not an
issue here: varying them changes the refined model parameters of
course, but they do not appear in the structure factor formula, so
don't need to be specified in the mathematical model to obtain the
Fcalcs.  You would of course need to know all the weights & target
values (as well as the SF formula) to reproduce the refinement to get
the deposited model.

But could future programs really re-calculate the same structure  
factors

from the deposited model? Because of the expected development of more
advanced methods and algorithms, I have my doubts ... *sigh*


Yes, if the deposited mathematical model is completely specified in
terms of the SF formula used and the values of *all* the parameters
that go into it, then in principle future versions of software using
more advanced models will be able to reproduce the exact Fcalcs.  This
assumes that the advanced models will use the same 'core' formula but
with additional terms and adjustable parameters, so that the simple
model can be obtained from the advanced one by constraining the extra
parameters to fixed values.  However if the simple model is not
'nested' inside the more advanced model in this way, then no it will
not be possible to reproduce the Fcalcs.

However as I implied, the main issue is that we're rather lax at fully
specifying our models (both formulae & parameters): obviously if in
future you don't have all the information you need to reproduce the
calculation then you have no hope of getting the same Fcalcs!

Cheers

-- Ian

Re: [ccp4bb] Deposition of riding H: R-factor is overrated

[Quyen Hoang]

Hi Nicholas,

Thank you for your reply.





it seems that we are trying to deposit one model to satisfy two
different purposes - one for model validation and the other for model
interpretation (use in docking etc), and what's good for one purpose
might not be necessarily good for the other.



This has been discussed before on this list, but allow me to repeat  
it:

You would have expected that the crystallographers' aim would be to
deposit the model that maximises the product (likelihood * prior).
Clearly, this is not what we do, mainly because (a) the calculation of
likelihood is only based on a subset of the 'data' that are obtained  
from
an X-ray diffraction experiment (for example, we ignore diffuse  
scattering
as Ian pointed-out), (b) we consciously avoid 'prior' because this  
would
make the models 'subjective', meaning that better informed people  
would
deposit (for the same data) different models than the less well  
informed,

(c) the format of the PDB does not offer much room for 'creative
interpretations' of the electron density maps [for example, you  
can't have
discrete disorder on the backbone (or has this changed ?)]. I sense  
that
what is being deposited is not the 'best model' in any conceivable  
way,
but the model that 'best' accounts for the final 2mFo-DFc map within  
the

limitations of the program used for the final refinement.


I don't quite understand your point. We currently deposit electron  
densities and movies, I don't see how depositing an energy minimized  
structure is so difficult. It doesn't need to be on the same pdb file  
as the model used in refinement nor does it need to be deposited into  
the PDB server, but even if it does, is it not possible to have it as  
a new "Chain" or new "atom type" in the current pdb file format?




ps. May I say parenthetically that making the deposited models  
dependant

on their intended usage, would possibly qualify as 'fraud' ;-)


I don't quite understand this either. When I prepare a protein model  
for simulation, I would remove all alternative conformations, add  
hydrogens, and then minimize the structure. If I make such a minimized  
structure available for others to use with full disclosure, how would  
that constitute "fraud"? I was going to start offering minimized  
models on our future structures on our lab website, but if that  
constitutes fraud, then I might have to rethink.


I don't know enough to argue with anyone here and that's not the  
intention of my posts - I am just trying to help figure out a way to  
resolve a significant problem that will likely to resurface down the  
road. It would be helpful if the more experienced people here can  
start a discussion of 'how to resolve' the problems exposed by this  
thread so far - assuming that you agree that it's a problem worth your  
time.


Cheers,
Quyen

__
Quyen Hoang, Ph.D
Assistant Professor
Department of Biochemistry and Molecular Biology,
Stark Neurosciences Research Institute
Indiana University School of Medicine
635 Barnhill Drive, Room MS0013D
Indianapolis, Indiana 46202-5122

Phone: 317-274-4371
Fax: 317-274-4686
email: qqho...@iupui.edu



--


 Dr Nicholas M. Glykos, Department of Molecular Biology
and Genetics, Democritus University of Thrace, University Campus,
 Dragana, 68100 Alexandroupolis, Greece, Tel/Fax (office)  
+302551030620,

   Ext.77620, Tel (lab) +302551030615, http://utopia.duth.gr/~glykos/

[ccp4bb] Postdoctoral Position in Structure Biology of Neurodegenerative Disease

[Quyen Hoang]

Postdoctoral position available:

A postdoctoral position is available to study the mechanisms of  
Parkinson disease. Genetic studies in the past decade have identified  
a number of genes as causal factors of Parkinson disease. Our goals  
are to understand the biochemical and biological functions of the  
disease associated proteins.


Our lab is located in the Department of Biochemistry and Molecular  
Biology at IUPUI (http://www.iupui.edu/) in Indianapolis Indiana  
(http://en.wikipedia.org/wiki/Indianapolis).


We are looking for an enthusiastic person who holds a PhD in  
biochemistry or related field, has a strong background in molecular  
biology and protein purification, and has strong interest in  
neuroscience.


Please email a cover letter, CV, and three reference letters to:
qqho...@gmail.com


Thank you for looking.

Cheers,
Quyen

Re: [ccp4bb] sulfur sad phasing

[Quyen Hoang]

I played around with a few years back - tried all kinds of different  
data collection strategies and wavelengths.
What worked for me was collecting data at wavelength of 1.7A from a  
few different crystals and then merge them together to get a higher  
redundancy.
Both calcium and sulfur signals were visible and provided enough  
phasing power.


Bone recognition mechanism of porcine osteocalcin from crystal  
structure.

Hoang QQ, Sicheri F, Howard AJ, Yang DS.
Nature. 2003 Oct 30;425(6961):977-80.

Radiation damage was a concern with longer wavelength.

Cheers,
Quyen

___
Quyen Hoang, Ph.D
Department of Biochemistry and Molecular Biology,
Start Neurosciences Research Institute
Indiana University School of Medicine
635 Barnhill Drive, Room MS0013D
Indianapolis, Indiana 46202-5122

Phone: 317-274-4371
Fax: 317-274-4686
email: qqho...@iupui.edu

On Nov 11, 2009, at 10:23 AM, Jim Pflugrath wrote:


2.29 Angstrom is an excellent wavelength for Cl, Ca and S anomalous
scatterers.  This is the wavelength produced by a Cr home source.
Some older references:

Acta Crystallogr D Biol Crystallogr. 2003 Nov;59(Pt 11):1943-57.  
Epub 2003
Oct 23. Away from the edge: SAD phasing from the sulfur anomalous  
signal

measured in-house with chromium radiation.

Acta Crystallogr D Biol Crystallogr. 2005 Jul;61(Pt 7):960-6. Epub  
2005 Jun
24. Away from the edge II: in-house Se-SAS phasing with chromium  
radiation.


Acta Crystallogr D Biol Crystallogr. 2005 Aug;61(Pt 8):1013-21. Epub  
2005

Jul 20. Structure determination of a novel protein by sulfur SAD using
chromium radiation in combination with a new crystal-mounting method.

I'm sure there are several structures done with Cr radiation since  
the above

articles were published.  Does any body else want to chime in?

Jim

-Original Message-
From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of
Matthias Zebisch
Sent: Wednesday, November 11, 2009 6:16 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] sulfur sad phasing

Dear bb!

What is the optimal wavelength for Sulfur SAD phasing?
Is it 1.9A or should one go below that to reduce absorption/damage.

Also, would the same wavelength be appropriate to maximize anomalous
scattering to position chlorides, calcium, sulfate in already phased
structures?

Thanks in advance,

Matthias

Re: [ccp4bb] Optimisation of 2D very thin plates to thick plates

[Quyen Hoang]

Hi Will,
We had a similar case recently.
Microseeding gave us single crystals (originally clusters), but still very thin 
plates as yours.
We then mutated a couple surface lysine residues to alanine and that led to 
thicker crystals.
The thin crystals were indexed as C2221, but the thicker ones revealed that it 
was actually P21.

Hope this helps.

Cheers,
Quyen

Quyen Hoang, PhD
Associate Professor of Biochemistry and Molecular Biology
Adjunct Associate Professor of Neurology
Primary Investigator of the Stark Neuroscience Research Institute
Indiana University School of Medicine
635 Barnhill Drive, MS0013C
Indianapolis, IN, 46202
(317)274-4371

> On Sep 19, 2019, at 12:16 PM, Phoebe A. Rice  wrote:
> 
> Is it a complex with DNA?  If so, vary the DNA ends?
>  
> ~~~
> Phoebe A. Rice
> Dept. of Biochem & Mol. Biol. and
>   Committee on Microbiology
> https://voices.uchicago.edu/phoebericelab/ 
> <https://voices.uchicago.edu/phoebericelab/>
>  
>  
> From: CCP4 bulletin board  <mailto:CCP4BB@JISCMAIL.AC.UK>> on behalf of William Richardson 
>  <mailto:william.richard...@nottingham.ac.uk>>
> Reply-To: William Richardson  <mailto:william.richard...@nottingham.ac.uk>>
> Date: Thursday, September 19, 2019 at 10:41 AM
> To: "CCP4BB@JISCMAIL.AC.UK <mailto:CCP4BB@JISCMAIL.AC.UK>" 
> mailto:CCP4BB@JISCMAIL.AC.UK>>
> Subject: [ccp4bb] Optimisation of 2D very thin plates to thick plates
>  
> Dear colleagues, 
> I would like some advice on crystal growth optimisation. I have been trying 
> to crystallise a DNA regulator. I got microcrystals in 1.3M Lithium Sulphate 
> and 50 mM Cacodylate pH = 6. Through a combination of condition refinement 
> and seeding I have attained crystals with a plate-like habit of dimension 
> 50micron, 50micron, <1micron. These have very weak (8Å) anisotropic 
> diffraction at DLS i24 but as they’re so wafer thin there has been no 
> improvement. Any suggestions to make them more 3D would be great. The space 
> group is C2221 
> Any advice?
> Thanks in advance
> Regards,
>  
> Will
>  
>  
> 
>  
> This message and any attachment are intended solely for the addressee
> and may contain confidential information. If you have received this
> message in error, please contact the sender and delete the email and
> attachment. 
>  
> Any views or opinions expressed by the author of this email do not
> necessarily reflect the views of the University of Nottingham. Email
> communications with the University of Nottingham may be monitored 
> where permitted by law.
>  
>  
>  
>  
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 
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To unsubscribe from the CCP4BB list, click the following link:
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[ccp4bb] Postdoctoral Positions Available

[Quyen Hoang]

Postdoctoral Positions at Indiana University School of Medicine 

My lab will have two postdoc positions available, one immediately and the other 
in April 2015. A summary of what we do can be found at 
http://snri.iusm.iu.edu/people/primary-investigators/quyen-q-hoang-ph-d/ 

No particular skill sets are required. We use a wide range of techniques 
including computational chemistry, molecular biology, protein biochemistry, 
enzymology, X-ray crystallography and other biophysical methods, yeast 
genetics, cell biology, and a little bit of C. elegans. I will show you these 
techniques as they become applicable to your project(s), but you must come with 
a desire to learn and a passion to understand the biological systems at the 
molecular level.

If you are interested, please email your CV and 3 reference letters to 
qqho...@iu.edu .

Cheers,

Quyen

Re: [ccp4bb] Off topic - Mercury Lamp for Akta

[Quyen Hoang]

We buy UV lamps from Jelight Company Inc. for replacements on our AKTA systems 
(http://www.jelight.com/low-pressure-uv-lamps.html 
).
It’s not a direct replacement in that a few modifications needed to be made:
1. cut two groves on the lamp base (with a file) to fit the GE housing.
2. needs it’s own power supply (12V DC), also available from Jelight.
The UV lamp costs $299.00, power supply costs $169.00.

The original GE lamps typically lasted between 1 to 2 years in my lab. The 
Jelight lamps have been running for more than 5 years and are still going.

Cons: the down side is that when the AKTA first starts up, it detects and 
adjust power output to the lamp, so we would need to turn the modified external 
12V DC power on at precisely the moment the AKTA looks for UV signals (by 
looking the status LED panel on the AKTA). Another minor drawback is that we 
bought 285nm lamps, so it can’t be switched to 260nm  for DNA (which we don’t 
need), but you can also buy a 260nm if you do need it.

Side note: besides the cost savings of the lamp, the lamp power supply in our 
AKTA burnt out (both units in my lab) and they cost $4,000. in parts alone to 
replace. The $169 external power supplies have been running constantly for 5 
years now without sign of problems.

Cheers,
Quyen

Assistant Professor
Indiana University School of Medicine
Department of Biochemistry and Molecular Biology
Stark Neurosciences Research Institute
635 Barnhill Drive
MS0013C
Indianapolis, IN 46202

> On May 6, 2015, at 8:48 AM, Mark Stead  wrote:
> 
> We have recently purchased a mercury lamp without the housing (product code: 
> 28-9354-93) from GE Healthcare.  The difference in cost between the two is 
> only small here in the UK, but I’m not sure how this compares to the States.
>  
> Mark
>  
>  
> Dr Mark A. Stead
> School of Biology | University of Leeds |
> L C Miall Building Room 8.16 | Leeds LS2 9JT | United Kingdom |
> 
> Lab: +44 (0)113 343 2890 | Office: +44 (0)113 343 3154 | E-mail: 
> m.a.st...@leeds.ac.uk 
> From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK 
> ] On Behalf Of Bonsor, Daniel
> Sent: 05 May 2015 16:06
> To: CCP4BB@JISCMAIL.AC.UK 
> Subject: [ccp4bb] Off topic - Mercury Lamp for Akta
>  
> The mercury lamp (28-4042-25) offered by GE Healthcare for Akta systems come 
> with the housing and is priced at $1345. We have plenty of sparing housing 
> units and are interested in just the lamp. Does anyone here have any cheaper 
> alternatives/suggestions than buying the lamp and housing?
>  
> Thanks in advance.
>  
> Dan
>  
> Daniel A Bonsor PhD.
> Sundberg Lab
> Institute of Human Virology
> University of Maryland, Baltimore
> 725 W Lombard Street N370
> Baltimore
> Maryland
> MD 21201
> Tel: (410) 706-7457
> http://www.sundberglab.org/Home.html 

Re: [ccp4bb] vitrification vs freezing

[Quyen Hoang]

I enjoyed following this thread. Because English is not my first language, I 
was hoping to learn the official definitions of these terms.
In my opinion, all the variations proposed so far are fine - I don't see 
problems with using them.

For me, when I see "flash frozen in liquid nitrogen" or "flash frozen in 
nitrogen stream" I get unambiguous mental images of how the crystals were 
prepared. When I hear a policeman yelling "freeze" while pointing a gun (no 
personal experience here), there is no ambiguity that I should stop moving (and 
won't get confused with cooling myself such that the water in my body would 
form hexagonal ice). When I hear that a person is frozen by Parkinson's 
disease, there is no ambiguity that his/her muscle had become rigid.

I think that I will continue to use "flash frozen in liquid nitrogen" or "flash 
frozen in nitrogen stream" and I hope that I would not need to explain to 
reviewers what that means.

Quyen



On Nov 16, 2012, at 10:48 AM, Ganesh Natrajan  wrote:

> Hi,
> 
> Maybe we  could just state the obvious,  ie, that the crystals were 
> 'Cryo-preserved' in liquid N2.
> 
> 
> Cheers
> 
> Ganesh
> 
> Le 16/11/12 16:27, Enrico Stura a écrit :
>> As a referee I also dislike the word "freezing" but only if improperly used:
>> "The crystals were frozen in LN2" is not acceptable because it is the outside
>> liquor that is rapidly cooled to cryogenic temperatures.
>> 
>> But the use of "freezing" used as the opposite of "melting" is fine and does 
>> not
>> imply a crystalline state. Ice is not always crystalline either:
>> http://en.wikipedia.org/wiki/Amorphous_ice
>> 
>> 

Re: [ccp4bb] vitrification vs freezing

[Quyen Hoang]

I was going to mention that too, but since I was a postdoc of Petsko my words 
could have been viewed as biased.

Quyen



On Nov 16, 2012, at 1:26 PM, Ronald E Stenkamp  
wrote:

> I'm a little confused.  Petsko and others were doing 
> low-temperature/freezing/vitrification crystal experiments in the 1970s, 
> right?  (J. Mol. Biol., 96(3) 381, 1975).  Is there a big difference between 
> what they were doing and what's done now.
> 
> Ron
> 
> On Fri, 16 Nov 2012, Gerard Bricogne wrote:
> 
>> Dear all,
>> 
>>I think we are perhaps being a little bit insular, or blinkered, in
>> this discussion. The breakthrough we are talking about, and don't know how
>> to call, first occurred not in crystallography but in electron microscopy,
>> in the hands of Jacques Dubochet at EMBL Heidelberg in the early 1980s (see
>> for instance http://www.unil.ch/dee/page53292.html). It made possible the
>> direct imaging of molecules in "vitrified" or "vitreous" ice and to achieve
>> higher resolution than the previous technique of negative staining. In that
>> context it is obvious that the vitreous state refers to water, not to the
>> macromolecular species embedded in it: the risk of a potential oxymoron in
>> the crystallographic case arises from trying to choose a single adjective to
>> qualify a two-component sample in which those components behave differently
>> under sudden cooling.
>> 
>>I have always found that an expression like "flash-frozen" has a lot
>> going for it: it means that the sample was cooled very quickly, so it
>> describes a process rather than a final state. The fact that this final
>> state preserves the crystalline arrangement of the macromolecule(s), but
>> causes the solvent to go into a vitreous phase, is just part of what every
>> competent reviewer of a crystallographic paper should know, and that ought
>> to avoid the kind of arguments that started this thread.
>> 
>> 
>>With best wishes,
>> 
>> Gerard.
>> 
>> --
>> On Thu, Nov 15, 2012 at 11:35:46PM -0700, Javier Gonzalez wrote:
>>> Hi Sebastiano,
>>> 
>>> I think the term "vitrified crystal" could be understood as a very nice
>>> oxymoron (http://www.oxymoronlist.com/), but it is essentially
>>> self-contradictory and not technically correct.
>>> 
>>> As Ethan said, "vitrify" means "turn into glass". Now, a glass state is a
>>> disordered solid state by definition, then it can't be a crystal. A
>>> vitrified crystal would be a crystal which has lost all three-dimensional
>>> ordering, pretty much like the material one gets when using the wrong
>>> "cryo-protectant".
>>> 
>>> What one usually does is to soak the crystal in a "cryo-protectant" and
>>> then flash-freeze the resulting material, hoping that the crystal structure
>>> will be preserved, while the rest remains disordered in a solid state
>>> (vitrified), so that it won't produce a diffraction pattern by itself, and
>>> will hold the crystal in a fixed position (very convenient for data
>>> collection).
>>> 
>>> Moreover, I would say that clarifying a material is vitrified when
>>> subjected to liquid N2 temperatures would be required only if you were
>>> working with some liquid solvent which might remain in the liquid phase at
>>> that temperature, instead of the usual solid disordered state, but this is
>>> never the case with protein crystals.
>>> 
>>> So, I vote for "frozen crystal".-
>>> 
>>> Javier
>>> 
>>> 
>>> PS: that comment by James Stroud "I forgot to mention that if any
>>> dictionary is an authority on the very cold, it would be the Penguin
>>> dictionary.", is hilarious, we need a "Like" button in the CCP4bb list!
>>> 
>>> --
>>> Javier M. Gonzalez
>>> Protein Crystallography Station
>>> Bioscience Division
>>> Los Alamos National Laboratory
>>> TA-43, Building 1, Room 172-G
>>> Mailstop M888
>>> Phone: (505) 667-9376
>>> 
>>> 
>>> On Thu, Nov 15, 2012 at 2:24 PM, Craig Bingman 
>>> wrote:
>>> 
 "cryopreserved"
 
 It says that the crystals were transferred to cryogenic temperatures in an
 attempt to increase their lifetime in the beam, and avoids all of the other
 problems with all of the other language described.
 
 I was really trying to stay out of this, because I understand what
 everyone means with all of their other word choices.
 
 On Nov 15, 2012, at 2:07 PM, James Stroud wrote:
 
> Isn't "cryo-cooled" redundant?
> 
> James
> 
> On Nov 15, 2012, at 11:34 AM, Phil Jeffrey wrote:
> 
>> Perhaps it's an artisan organic locavore fruit cake.
>> 
>> Either way, your *crystal* is not vitrified.  The solvent in your
 crystal might be glassy but your protein better still hold crystalline
 order (cf. ice) or you've wasted your time.
>> 
>> Ergo, "cryo-cooled" is the description to use.
>> 
>> Phil Jeffrey
>> Princeton
>> 
>> On 11/15/12 1:14 PM, Nukri Sanishvili wrote:
>>> s: An alternative way to avoid the argument and discussion a

Re: [ccp4bb] vitrification vs freezing

[Quyen Hoang]

Hi Ed,

> If we speak the way scientific articles are written...
> 
> By Bernard Dixon, published in New Scientist, 11 April 1968, p.73, an 
> imaginary conversation at breakfast:
> 
> "Daddy, I want cornflakes this morning. Must I have porridge?"
> 
> "Yes. It has been suggested by mummy that, in view of the external coldness, 
> the eating of porridge by you will cause an increase in bodily temperature. 
> Furthermore, in regard to the already-mentioned temperature considerations, 
> your grandma-knitted gloves and wool-lining-hooded coat will have to be worn."

Translating into common language: 
"Yes. It's cold outside, having porridge will keep you warmer and please wear 
your gloves and wool coat."

> 
> "May I have some sugar on my porridge?"
> 
> "The absence of sugar in the relevant bowl has been noted by daddy at an 
> earlier moment. However, further supplies of this substance are now being 
> brought by mummy from the appropriate vessel that is present in the kitchen."
> 

Translating into common language: 
"The sugar bowl was empty, but mommy had replenished it."

If scientific articles are written as long winded and ambiguous as above 
examples, which could be easily made more concise and less ambiguous in common 
language (as suggested above), then I favor reporting scientific findings using 
common language.

BTW, Ed, I have been hoping that F1 would return to Indy so that you guys would 
come to visit ;)

Quyen

> 
> 
> -- 
> Oh, suddenly throwing a giraffe into a volcano to make water is crazy?
>Julian, King of Lemurs

Re: [ccp4bb] Contagious, Self-Distributing "Vaccines?"

[Quyen Hoang]

 I guess that one might need a way to avoid natural mutations that could turn B 
into B’ or B".

Quyen

Quyen Hoang, PhD
Associate Professor of Biochemistry and Molecular Biology
Adjunct Associate Professor of Neurology
Primary Investigator of the Stark Neuroscience Research Institute
Indiana University School of Medicine
635 Barnhill Drive, MS0013C
Indianapolis, IN, 46202
(317)274-4371
https://qqhoang.pages.iu.edu

>> It would seem to me that it should be possible to generate versions of the 
>> Covid virus that would:
>> 
>> A. be extremely contagious and yet
>> B. be clinically benign, and
>> C. confer immunity to the original covid virus.
>> 
>> If, then, this virus could be released, with appropriate "kill switch" 
>> safeguards built in, would this not solve the world's pandemic problems? Is 
>> there any reason, practically, why this approach would not be feasible?
>> 
>> Maybe we don't really know enough to manipulate A, B, C yet?
>> 
>> Or maybe it's too scary for primetime...nightmare bio-warfare apocalypse?
>> 
>> Has this sort of thing been done, or does it have a name?
>> 
>> Jacob
>> -- 
>> +
>> 
>> Jacob Pearson Keller
>> 
>> Assistant Professor
>> 
>> Department of Pharmacology and Molecular Therapeutics
>> 
>> Uniformed Services University
>> 
>> 4301 Jones Bridge Road
>> 
>> Bethesda MD 20814
>> 
>> jacob.kel...@usuhs.edu <mailto:jacob.kel...@usuhs.edu>; 
>> jacobpkel...@gmail.com <mailto:jacobpkel...@gmail.com>
>> Cell: (301)592-7004
>> 
>> +
>> 
>> 
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 
>> <https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1>
> -- 
> ===
> All Things Serve the Beam
> ===
>David J. Schuller
>modern man in a post-modern world
>MacCHESS, Cornell University
>schul...@cornell.edu 
> <mailto:schul...@cornell.edu>
> To unsubscribe from the CCP4BB list, click the following link:
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[ccp4bb] Postdoc positions

[Quyen Hoang]

Hi All,

My group has a couple of Postdoc positions available immediately to study the 
structures and functions of proteins associated with neurodegenerative diseases.
The projects utilize cryo-EM and X-ray crystallography along with a battery of 
other biophysical and biochemical methods.
As such, they might be good opportunities for crystallographers looking to 
learn cryo-EM or microscopists wishing to learn X-ray crystallography (however, 
experiences with these methods are not necessary to apply).
These positions are supported by a new 5-year NIH grant.

Please email me directly (qqho...@iu.edu) if you are interested in learning 
more about these positions.

Cheers,
Quyen

___
Quyen Hoang, PhD
Associate Professor of Biochemistry and Molecular Biology
Director of IUSM Center for Electron Microscopy (iCEM)
Adjunct Associate Professor of Neurology
Primary Investigator of the Stark Neuroscience Research Institute
Indiana University School of Medicine
635 Barnhill Drive, MS0013C
Indianapolis, IN, 46202
(317)274-4371
https://qqhoang.pages.iu.edu



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[ccp4bb] Postdoctoral position in structural biology of neurodegeneration

[Quyen Hoang]

Hi All,
We have a postdoctoral position available and would appreciate your help in 
spreading the word.

Postdoctoral Position in Structural and Chemical Biology of Neurodegenerative 
diseases.

Position Description: Applicants are invited to apply for a postdoctoral 
fellowship position in Dr. Hoang's laboratory at Indiana University School of 
Medicine.  Our research focuses on delineating the molecular mechanisms of 
neurodegeneration. We use a broad range of experimental methods, including 
molecular dynamic simulation and in silico docking, molecular cloning, protein 
biochemistry, enzymology, X-ray crystallography, electron microscopy (including 
cryo-EM), NMR, yeast genetics, mammalian cell models, and C. elgans. Basically, 
we would go wherever the science takes us. Examples of some the methods that we 
employ can be gleaned from our prior: Hoang QQ et al. Nature. 425: 977-980 
(2003); Liao J. et al. PNAS. 111(11):4055-60 (2014), Oh M et al. PNAS. 
111(30):11007-12 (2014), and Wang W. et al. PNAS. 113(34):9587-92 (2016).

Requirements: We expect this position to be filled with a recent PhD in 
biophysics, biochemistry, chemistry, bioengineering, or a related discipline 
providing solid knowledge of biological macromolecules.  Experience in protein 
expression and purification is essential, as is familiarity with 
thermodynamics, biochemistry, and quantitative data analysis.  We are 
particularly interested in a person who has expertise in one or more methods 
mentioned above and is looking to learn new complimentary skills to carry out a 
multidisciplinary investigation into molecular mechanisms of brain 
degeneration. The applicant needs to be a team worker with good communication 
skills.

To Apply:
Applicants should submit the following: brief description of research 
interests and career goals, CV, and contact information for three references to 
Dr. Quyen Hoang (qqho...@iu.edu).


Cheers,
Quyen

Quyen Hoang, PhD
Associate Professor of Biochemistry and Molecular Biology
Adjunct Associate Professor of Neurology
Primary Investigator of the Stark Neuroscience Research Institute
Indiana University School of Medicine
635 Barnhill Drive, MS0013C
Indianapolis, IN, 46202
(317)274-4371

Re: [ccp4bb] To Trim or Not to To Trim

[Quyen Hoang]

As with Jurgen, I’ve never trimmed a residue.
In my view the trimmed residues do not exist.
If we were to build models consisting of only atoms defined by the experimental 
density, then I wonder what the original model of the DNA double helix would 
have looked like.

BTW, hey Jurgen, long time no talk.

Cheers,
Quyen

Quyen Hoang, PhD
Professor of Biochemistry and Molecular Biology
Director of IUSM Center for Electron Microscopy (iCEM)
Adjunct Professor of Neurology
Primary Investigator of the Stark Neuroscience Research Institute
Indiana University School of Medicine
635 Barnhill Drive, MS0013C
Indianapolis, IN, 46202
(317)274-4371
https://qqhoang.pages.iu.edu


> On Mar 10, 2023, at 12:09 PM, Jurgen Bosch  wrote:
> 
> I’m sure James H. Is preparing a philosophical dissertation on the "State of 
> the atoms to B or not to B that is not only a refinement question” that he 
> will share momentarily with the board.
> 
> Jürgen 
> 
>> On Mar 10, 2023, at 12:06 PM, DEBANU DAS  
>> wrote:
>> 
>> Yes, zero occupancy would reflect that. But not the coordinates in any 
>> proper way. 
>> Debanu
>> 
>> On Fri, Mar 10, 2023 at 8:58 AM Jurgen Bosch > <mailto:jxb...@case.edu>> wrote:
>> Going back to RIP phasing methods :-)
>> So Harry in your particular case occupancy of zero would actually reflect 
>> reality for those “combusted” atoms.
>> 
>> Jürgen 
>> 
>> > On Mar 10, 2023, at 11:56 AM, Harry Powell 
>> > <193323b1e616-dmarc-requ...@jiscmail.ac.uk 
>> > <mailto:193323b1e616-dmarc-requ...@jiscmail.ac.uk>> wrote:
>> > 
>> > Hi folks
>> > 
>> > One other thing that I haven’t noticed anyone mentioning yet (sorry to 
>> > those who have mentioned it!!) is that you may not see your sidechain 
>> > atoms in density because they are not there at all, in spite of what you 
>> > may have had in the original protein, or even if the atoms were really 
>> > there in the crystal _before_ exposure to the beam.
>> > 
>> > The coordinates are supposed to be what you actually find, not what you 
>> > hope is there.
>> > 
>> > Just my two ha’porth
>> > 
>> > Harry
>> > 
>> > 
>> > 
>> > To unsubscribe from the CCP4BB list, click the following link:
>> > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 
>> > <https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1>
>> > 
>> > This message was issued to members of www.jiscmail.ac.uk/CCP4BB 
>> > <http://www.jiscmail.ac.uk/CCP4BB>, a mailing list hosted by 
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>> 
>> 
>> 
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> 
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Re: [ccp4bb] To Trim or Not to To Trim

[Quyen Hoang]

Hi Debanu,In your example, I would guess that models with truncated side chains would probably be problematic too, so would alternative conformations, unmodelled segments, asymmetric unit consisting of only a fraction of the biological/functional unit, etc.I am not sure if it is possible to build a model in a way that would avoid all the potential pitfalls in the circumstance you mentioned. Or should one even try?I don't remember ever solving a structure for the purpose of providing a model for computational chemists to use. We solve structures to answer certain questions and I would rather deposit the models we used to gain the insight we seeked instead of the one artificially made in the attempt to avoid misuses or understanding by non-structural biologists. Cheers,QuyenOn Mar 10, 2023, at 4:01 PM, Debanu Das  wrote:Hi Tom,-"so no matter how much we try to educate people, the vast (vast, vast) majority of people will take
 the models as the 'truth'-"So if we don't see something, the conservative approach is to probably 
avoid putting it in, otherwise it will get propagated forever"I absolutely agree. I have a recent anecdote about this to share. I was at a workshop where the majority of attendees were medicinal chemists, comp chemists, biologists. I found out that some (many?) were even performing comp chem studies (where ligands and waters were important) on structures downloaded straight from the PDB without even checking associated electron density for such ligands/waters or even realizing potential side chain issues (not even including about any potential main chain or other quality/validation issues).   Best regards,Debanu--Debanu Das,https://bio.site/debanu_dasOn Fri, Mar 10, 2023 at 12:41 PM Tom Peat  wrote:






Hello All, 




I agree with Dale that we don't have a good way to model these things and it has been a discussion for a very long time with no proper answer. 




Two more small points- we (or at least I do this all the time) don't model the N- or C-terminal residues that I don't see. They are most likely there somewhere (waving goodbye in the solvent mask) and that seems to be (relatively) standard practice, so I'm
 not sure how different this is to not putting in side chains that can't be seen? We don't replace with Ala, so people should know what the actual sequence is, but there is no evidence for a side chain. A multi-model is likely the better way to go, but isn't
 as feasible currently as the single model we normally deposit. As mentioned by others, we shouldn't try to put in ligands or co-factors that we don't see either... 




One other point- although we should educate, the PDB has estimated that 99% of the users are not structural biologists (nor modellers or others with training), so no matter how much we try to educate people, the vast (vast, vast) majority of people will take
 the models as the 'truth'. So if we don't see something, the conservative approach is to probably avoid putting it in, otherwise it will get propagated forever. 




My two cents. cheers, tom 


From: CCP4 bulletin board  on behalf of Dale Tronrud 
Sent: Saturday, March 11, 2023 7:15 AM
To: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] To Trim or Not to To Trim
 


Hi

    As a frequent contributor to prior discussions on this same topic I 
would like to broaden the discussion a bit.  I'm sorry to say that most 
of the comments on this threat are exactly the same positions that have 
been expressed many times over the years.  I don't want to spent time, 
again, retyping my opinions on how I prefer to torment the parameters of 
my models to express what I believe is going on inside my crystals.

    The fundamental problem is that the parameters we are forced to use, 
in our PDB depositions and in the refinement and model building programs 
available to us, are wholly inadequate.  We cannot accurately (or 
precisely) describe what we are are envisioning for the surface side 
chains, and sometimes entire stretches of main chain, in our proteins. 
We can continue to argue with each other year after year, but there is 
no solution to this problem other than changing the nature of PDB models 
and allowing a reasonable description of multi-conformation models.

    I believe it is fair to say that the consensus after a previous 
round of this discussion was that, at the very least, we need a flag for 
each atom which indicates whether that atom was placed based on electron 
density or simply to make a chemically complete set of atoms for that 
type of monomer.  I haven't looked but I think that was about five or 
ten years ago.  Since then the PDB has made major changes to the 
structure of PDB entries that will require most software for analysis of 
macromolecular models be rewritten and right now that organization is 
making a major push to get us to virtually attend a workshop to help us 
make this transition.  And

Re: [ccp4bb] [External] Re: [ccp4bb] Structure prediction - waiting to happen

[Quyen Hoang]

Thank you for having this conversion. I have heard this question more than once (why is experimental determination needed when there is Alphafold) at grant study sections. Some came from structural biologists, so hopefully, the discussion here would help them also. Computational modeling had existed for decades before Alphafold, and no one ever questioned then, as far as I am aware, the need for experimental determination of the modeled structures regardless of how accurate the predicted models may have been (for example an MD state of a point mutation using a high-resolution X-ray model as a starting model), so I am confused as to why we are asking this question now. In my view, predicted model, as good as it may be, is still a prediction. Cheers,QuyenQuyen Hoang, PhDAssociate Professor Department of Biochemistry and Molecular Biology Adjunct Associate Professor of NeurologyPrincipal Investigator of the Stark Neurosciences Research InstituteIndiana University School of Medicine635 Barnhill DriveMedical Sciences Building, room MS0013CIndianapolis, IN 46202317-274-4371On Apr 2, 2023, at 10:52 AM, Eugene Valkov  wrote:It depends on the problem under investigation. For example, if you propose to perform drug discovery with membrane proteins in a lipid/detergent environment or study the structural organization of large ribonucleoprotein assemblies, then AlphaFold will be of limited value and you should point this out in the rebuttal.If one of your aims is to study a protein, divide it into pieces, and solve structures of domains, then this is justifiably questioned by reviewers in light of the information provided by structure prediction. Perhaps in the reviewer's opinion, there is no need to use substantial funds and time to solve that structure. A more competitive proposal may then re-focus on requesting resources for using the structural hypotheses to probe the mechanism with more ambitious experimental structural aims or to take a deeper dive into the function.As Darwin said, it is not the strongest or the fastest (or even the most intellectual!) that survives.On Sun, 2 Apr 2023 at 11:28, Srivastava, Dhiraj  wrote:




That’s the point. All these predictions can not replace experiments and they should be used only in the absence of experimental structures, that too with caution. A biophysicist and structural biologist understand this but most of the non-experts (including
 the reviewers of the grants from non-structural biology study sections) don’t understand this and think that with alphafold, there is no need for experimentally determined structures. That’s more damaging than helpful. 




From: CCP4 bulletin board  on behalf of Eugene Valkov 
Sent: Sunday, April 2, 2023 10:10 AM
To: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] [External] Re: [ccp4bb] Structure prediction - waiting to happen
 



Predicted
 structures lack the precision and accuracy of experimentally-determined structures at high resolution with all the benefits of unbiased co-discovery of solvent molecules, ions, ligands, etc., bound to molecules of interest.

It
 is also true that AI-assisted structure prediction can be stunningly accurate to the level of correctly identifying interfacing residues between interacting proteins and accurately recapitulating the architectures of large, multi-domain complexes. AI-assisted
 structure prediction is immensely liberating in lowering the threshold to generate testable hypotheses for those lacking access to experimental structural biology resources.

AlphaFold
 and related structure prediction tools are here to stay, and no magic incantations in funding proposals will likely make them disappear. So rather than wring our hands as a community and mourning the loss of ‘divide-and-conquer’ structural biology, which was
 a rich vein to mine over the last decades, we should adapt to structure prediction and normalize its use, with proper controls and caveats, as part of our arsenal of tools and methods to focus on the questions under investigation.

With
 best wishes,
Eugene

Eugene
 Valkov, D.Phil.
Stadtman
 Investigator
RNA
 Biology Laboratory
Center
 for Cancer Research
National
 Cancer Institute
Frederick
 MD 21702, USA
(301)
 846-1823




On Sun, 2 Apr 2023 at 10:44, 
jacinto.ls  wrote:



I am also not sure whether AlphaFold can address the impact of ions and other cofactors on the fold of many proteins.

Best wishes,
Jacinto
On 2/4/23 16:20, Srivastava, Dhiraj wrote:

May be this article is of some help suggesting the need of experimental structures despite excellent alphafold model.
https://www.nature.com/articles/s41401-022-00938-y



From: CCP4 bulletin board
 on behalf of Ian Tickle

Sent: Sunday, April 2, 2023 8:28 AM
To: CCP4BB@JISCMAIL.AC.UK

Subject: [External] Re: [ccp4bb] Structure prediction - waiting to happen
 





All, the first hurdle will of course be

[ccp4bb] Structure prediction - waiting to happen

[Quyen Hoang]

Hi River,

What's in a name? That which we call a predicted structure
By any other name would still be uncertain until validated experimentally

Cheers,
Quyen

> On Apr 3, 2023, at 2:12 PM, Xiao, Chuan  wrote:
> 
> You raised a very good question: why we are asking this question now? Maybe 
> the answer is somewhere on the fact that Alphafold is a “revolution” in the 
> prediction field (borrow the term from resolution revolution in cryo-EM).
>  
> Anyway, when reading all these discussions, it made me think how I came to US 
> to learn how to solve a structure. My Master training is more like 
> bioinformatics. My second paper has a component of structural homology 
> modeling after sequencing some Rice genes. The paper was rejected by the 
> reviewers asking how I could proof my structural modeling (prediction) was 
> correct. At that time, I decided to learn how to solve the structure to proof 
> my prediction is correct. Now, it is interesting to see now the question is 
> on the other side. 😊
>  
> River  
>  
> From: CCP4 bulletin board  <mailto:CCP4BB@JISCMAIL.AC.UK>> On Behalf Of Quyen Hoang
> Sent: Sunday, April 2, 2023 10:20 AM
> To: CCP4BB@JISCMAIL.AC.UK <mailto:CCP4BB@JISCMAIL.AC.UK>
> Subject: Re: [ccp4bb] [External] Re: [ccp4bb] Structure prediction - waiting 
> to happen
>  
> EXTERNAL EMAIL: This e-mail is from a sender outside of the UTEP system. Do 
> not click any links or open any attachments unless you trust the sender and 
> know the content is safe.
> Please forward suspicious emails to secur...@utep.edu 
> <mailto:secur...@utep.edu> or call 915.747.6324
> 
> Thank you for having this conversion.  
> I have heard this question more than once (why is experimental determination 
> needed when there is Alphafold) at grant study sections. Some came from 
> structural biologists, so hopefully, the discussion here would help them 
> also. 
> Computational modeling had existed for decades before Alphafold, and no one 
> ever questioned then, as far as I am aware, the need for experimental 
> determination of the modeled structures regardless of how accurate the 
> predicted models may have been (for example an MD state of a point mutation 
> using a high-resolution X-ray model as a starting model), so I am confused as 
> to why we are asking this question now. In my view, predicted model, as good 
> as it may be, is still a prediction. 
>  
> Cheers,
> Quyen
> 
> Quyen Hoang, PhD
> Associate Professor 
> Department of Biochemistry and Molecular Biology 
> Adjunct Associate Professor of Neurology
> Principal Investigator of the Stark Neurosciences Research Institute
> Indiana University School of Medicine
> 635 Barnhill Drive
> Medical Sciences Building, room MS0013C
> Indianapolis, IN 46202
> 317-274-4371 
>  
> 
> 
> On Apr 2, 2023, at 10:52 AM, Eugene Valkov  <mailto:eugene.val...@gmail.com>> wrote:
> 
>  
> It depends on the problem under investigation. For example, if you propose to 
> perform drug discovery with membrane proteins in a lipid/detergent 
> environment or study the structural organization of large ribonucleoprotein 
> assemblies, then AlphaFold will be of limited value and you should point this 
> out in the rebuttal.
>  
> If one of your aims is to study a protein, divide it into pieces, and solve 
> structures of domains, then this is justifiably questioned by reviewers in 
> light of the information provided by structure prediction. Perhaps in the 
> reviewer's opinion, there is no need to use substantial funds and time to 
> solve that structure. A more competitive proposal may then re-focus on 
> requesting resources for using the structural hypotheses to probe the 
> mechanism with more ambitious experimental structural aims or to take a 
> deeper dive into the function.
>  
> As Darwin said, it is not the strongest or the fastest (or even the most 
> intellectual!) that survives.
>  
> On Sun, 2 Apr 2023 at 11:28, Srivastava, Dhiraj  <mailto:dhiraj-srivast...@uiowa.edu>> wrote:
> That’s the point. All these predictions can not replace experiments and they 
> should be used only in the absence of experimental structures, that too with 
> caution. A biophysicist and structural biologist understand this but most of 
> the non-experts (including the reviewers of the grants from non-structural 
> biology study sections) don’t understand this and think that with alphafold, 
> there is no need for experimentally determined structures. That’s more 
> damaging than helpful. 
>  
>  
> From: CCP4 bulletin board  <mailto:CCP4BB@JISCMAIL.AC.UK>> on behalf of Eugene Valkov 
> mailto:eugene.val...@gmail.com>>
> Sent: Sunday, April 2, 2023 10

[ccp4bb] Technician Position available

[Quyen Hoang]

Hi All,

We have a technician position open with the possibility of transitioning to a 
PhD student in the lab.
I appreciate your help in spreading the word.

Technician Position in Structural and Chemical Biology of Neurodegenerative 
diseases.

Position Description: Applicants are invited to apply for a technician position 
in Dr. Hoang's laboratory in the Department of Biochemistry and Molecular 
Biology at Indiana University School of Medicine. Our research focuses on 
understanding the molecular mechanisms of neurodegeneration. We use a broad 
range of experimental methods, including molecular dynamic simulation and in 
silico docking, molecular cloning, protein biochemistry, enzymology, X-ray 
crystallography, cryo-electron microscopy (cryo-EM), NMR, yeast genetics, 
mammalian cell models, and C. elgans. Examples of some of the methods that we 
employ can be gleaned from our prior publications: Hoang QQ et al. Nature. 425: 
977-980 (2003); Liao J. et al. PNAS. 111(11):4055-60 (2014), Oh M et al. PNAS. 
111(30):11007-12 (2014), Wang W. et al. PNAS. 113(34):9587-92 (2016), Wu CX et 
al. JBC. 294(15):5907-5913(2019), and Wu CX et al. bioRxiv, 676627

Requirements: BS or MS in biophysics, biochemistry, chemistry, bioengineering, 
biology, or a related discipline. No experience is required. However, 
experience in protein expression and purification is a plus.

To Apply:
Please email your letter of interest, CV, and contact information for 
three references to Dr. Hoang (qqho...@iu.edu).

Cheers,
Quyen
__
Quyen Hoang, PhD
Professor of Biochemistry and Molecular Biology
Director of IUSM Center for Electron Microscopy (iCEM)
Adjunct Professor of Neurology
Primary Investigator of the Stark Neuroscience Research Institute
Indiana University School of Medicine
635 Barnhill Drive, MS0013C
Indianapolis, IN, 46202
(317)274-4371
https://qqhoang.pages.iu.edu




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