Hi,
Since my mailbox is swimming in 'Images!' emails I would add my irrelevant
two cents:
Image storage does not pay for itself. There has to be a source of funding
for it. Storing, transmitting, etc. of the ever-increasing number of
terabytes costs money, which at the moment no one seems to
Hello,
The short answer is 'yes'. If you can use both methods :) The issue with
limited proteolysis lies in the questionable state of the full-length
protein - if the stuff is nasty and misfolded, then fagments generated by
proteolytic digest aren't going to be meaningful. On the other hand if
Hi,
I wouldn't call this a standard procedure - generally speaking the most
important two parameters you have to consider are: the protein/ligand
affinity and the supply of the ligand. For tightly bound ligands, you may be
able to just add the ligand once (i.e. during cell growth and
Look up a standard endotoxin assay (usually an immunoassay) because that's
exactly what endotoxin is :-)
Artem
---
When the Weasel comes to give New Year's greetings to the Chickens no good
intentions are in his mind.
_
From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On
intentions are in his mind.
_
From: Artem Evdokimov [mailto:ar...@xtals.org]
Sent: Thursday, February 26, 2009 6:57 PM
To: 'JOE CRYSTAL'
Cc: 'CCP4BB@JISCMAIL.AC.UK'
Subject: RE: [ccp4bb] Questions regarding Peptidoglycan-binding protein
Look up a standard endotoxin assay (usually
The protein wolves get it - and it's never heard from or seen again.
Artem
---
When the Weasel comes to give New Year's greetings to the Chickens no good
intentions are in his mind.
-Original Message-
From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of Jacob
Keller
Unfortunately the database contains huge bias towards:
a) proteins that are easy to make and crystallize
b) biologically interesting protein families
Artem
---
When the Weasel comes to give New Year's greetings to the Chickens no good
intentions are in his mind.
-Original Message-
If it smells of octanol (vaguely fruity, cloying odor) and possibly has a
little phase separation (octanol poorly mixes with water) then you have
significant hydrolysis.
Beyond the smell test, you can run a small sample out on a TLC with
n-Octanol as control lane. Pretty easy.
As to
http://www.sciencedirect.com/science?_ob=ArticleURL_udi=B6V5N-4C8PFBS-2_us
er=10_rdoc=1_fmt=_orig=search_sort=dview=c_acct=C50221_version=1
_urlVersion=0_userid=10md5=aee9aceae8dfa17b363ee2ec634debb0
Osmotic shock is OK, or you can try chlorophorm disruption. You can Google
for either to find
Hi,
I had this exact problem before. The method below worked for me:
Take a sturdy needle (like one of the microneedles from a Hampton kit or a
very thin syringe needle) and (while observing the whole thing under a
scope) stick the needle into the plastic a bit away from the crystal. Push
Hi,
Yes, this does happen.
Spontaneous α-N-6-Phosphogluconoylation of a His Tag inEscherichia
coli:The Cause of Extra Mass of 258 or 178 Da in Fusion Proteins
http://www.sciencedirect.com/science?_ob=ArticleURL_udi=B6W9V-45N4K22-R_us
Hi,
A few simple hints:
(Please note that I am aware of the inexact language in the statements below
but I don't have the time to write this up exactly - conversational English
would have to do. Caveat emptor.)
Most protein crystals will break or deform when poked with a steel needle.
With respect to the surface entropy reduction method - if you have the
option of sharing your sequence, I can run it through some of my routines
too. Sorry - they're not available as public servers yet because they are
still in development. It significantly helps to have even a very low
resolution
Hi,
It sometimes comes down to manual dexterity - do you have a friend with
'surgical hands'? It's sometimes possible to simply cut a little 'window' in
the skin and to harvest the crystal, skin and all - usually the skin is not
significantly ordered and thus does not pose a problem for data
Is your phosphate a Na-K phosphate? If it is, then you're probably getting a
K salt of SDS which is considerably less soluble than Na or Li salts.
Artem
-Original Message-
From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of
Subscribe Ccp4Bb Kn L
Sent: Wednesday,
Interestingly, in the interactive 3D applet view of the ligand from the PDB
the two are perfectly in plane, whereas in the protein viewer the two groups
are clearly out of plane. I assume that this means that the coordinates for
the 3D ligand view are re-computed internally and are not
Folks,
This discussion is now dangerously close to a philosophical discourse
regarding the differences between homoplasy, homology, and analogy. Throw
into the mix synapomorphy and symplesiomorphy - and we've got ourselves a
cladistic analysis soup sprinkled with the croutons of phylogeny.
I do
It helps to remember that PCR does have an upper limit of total
double-stranded DNA content (regardless of its molarity!) after which it
does not work any more (due to the competition of the polymerase for
non-specific dsDNA versus primer-substrate pairs).
Therefore the theoretical limits on this
Hi,
I didn't want to get into this because so many good comments and suggestions
have already been made - but I guess I will make a wordy and effusive
comment of my own :)
In my opinion, the question of how *specific* tags (His, c-Myc) influence
crystallization is somewhat misleading. The
There are quite a few MBP fusions in the PDB. Just search using MBP sequence
and you will get (among others):
1A7L
1HSJ
1IUD
1MG1
1MH3
1NMU
1R6Z
And so on...
Artem
-Original Message-
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of David
M Shechner
Sent: Thursday,
Jacob,
We routinely express SeMet proteins in native E. coli without any trouble.
The method is a derivative of Doublie's. If you want, I would be more than
happy to send you a protocol (we've modified the process to gain biomass
yield).
Artem
-Original Message-
From: CCP4 bulletin
http://www.xtals.org/pdfs/selenium.pdf
(sorry, for some reason first email went out w/o the hyperlink)
Artem
-Original Message-
From: Artem Evdokimov [mailto:[EMAIL PROTECTED]
Sent: Monday, November 03, 2008 9:26 PM
To: 'Jacob Keller'; 'CCP4BB@JISCMAIL.AC.UK'
Subject: RE: [ccp4bb
Why soak for a whole minute? A single pass through cryo is usually enough,
and that takes a couple of seconds with the right set-up...
You could try oil - if you're lucky it solves your issues. Note that not all
oils are the same, and many people succeed with blended compositions rather
than pure
Tiancen,
While not entirely impossible - this is a formidable task. The answer to
your quesiton depends on the specifics of your situation and on what
additional information you are able to procure. Quite a few kinases have
specific sites; equal or greater numbers are only partially specific
If you're 100% sure that this is only one atom then amination comes to mind.
I have no clue what conditions would favor such reactivity but it is
possible that the formyl group on the Met was aminated with the cyclic N of
the histidine, resulting in either a substituted bis-amine (requires
Maybe we're not talking about the same kind of container? We never had any
kind of particles** in ours and with the use of a lid the ice does not
appear for quite some time. We did not see 'vigorous boiling' either. If one
leaves LN2 in the open it would eventually ice up regardless of the nature
Dear Meg,
Please note that there are many ways to do HIC. Different resins result in
different degrees of separation and it's unfortunately impossible to predict
what would happen without trying it out. Which is why various companies sell
HIC 'trial kits' - a few little columns packed with media
Dear Sue,
This is a very interesting case! Normally the Zn-S bond is quite strong - so
it's an unusual situation to have. Did you already attempt to purify the
protein in the presence of tiny quantities of Zn? Also, what buffers and
other components are you using during purification, and what is
Please note that TCEP decomposes and one of the decomposition products is
phosphate. Enough TCEP and you might have Zinc Phosphate crystals which can
sometimes look very odd and protein looking.
Artem
_
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of
Jennifer
Hi,
In addition to the excellend comments posted already, I would like to
venture an opinion regarding learning, technology, and associated matters.
Do not make the problem fit the techniques you know - instead try to make
sure that you know (or know where to learn) enough techniques to
Dear Matt,
Based on the follow-up information that you posted, I would be willing to
bet that the color you observe is caused by various Ni complexes formed with
DTT/BME/etc. Dialysis won't necessarily remove these complexes (especially
if protein is part of the complex) and EDTA may not destroy
Hi,
First of all - I am curious why did you decide to put in an extra step (the
T/A cloning into an intermediate vector)? You can happily digest your PCR
product with NheI/BamHI, clean up and ligate into the appropriately digested
pET-23a(+). If you have issues, you should definitely try
T/A cloning utilizes the overhangs left by certain polymerases as cloning
handles. To lower background, the vectors for TA cloning are often designed
to contain a rare cutter which is used during ligation to constantly re-cut
the self-ligated vector, so this way only the insert ligation product is
Dear ccp4ers,
Does someone by any chance have a snippet of code (in whatever language but
PERL, C, or FORTRAN for preference) that would perform coordinate fit for
two sets of atoms? I don't need anything fancy, just some simple code to fit
two peptides of equal length and composition. Yes, I
Thank you,
Courtesy of Peter Zwart, William Scott, Phil Evans, and Gerard Kleywegt - I
now have *four* different ways to do what I need! I can use Clipper, cctbx,
Fortran+numerical recipes, or Clipper :-)
This goes to show that crystallographers are:
a) overall very nice and
b)
Exactly.
There are about fifteen reasons that I could name without thinking as to why
any particular step in restriction-based cloning may not work. If I think
about it the number goes up to thirty or more. Yet only ONE of them is what
actually plagues you. So please Vijay, give us sufficient
exchanger.
Does Imidazole had any bad effect in protein. I have experienced that the
protein cannot sustain low salt and get ppt during dialysis.
Sincerely
Debajyoti Dutta
On Sat, 23 Aug 2008 Artem Evdokimov wrote :
Hi,
If you are plagued by 'generic' proteolysis, it is not likely that changing
Also, for very large proteins you should not boil the samples - if you must
heat, just go to 50-60C for a few minutes. You also may have better luck
with CTAB-PAGE rather than SDS-PAGE (CTAB tends to smear less, for some
reason).
Artem
-Original Message-
From: CCP4 bulletin board
Hi,
If you are plagued by 'generic' proteolysis, it is not likely that changing
buffers from TRIS to phosphate will help reduce the breakdown. You may want
to ask yourself several key questions regarding the breakdown:
1. is it proteolysis or abortive translation?
2. is it
As a follow-up:
About a week or two ago, SOLOMON worked very well at 2.4A Hg-SAD (1A) phased
map for ~1100 amino acids and 6 mercury sites. Almost 70% solvent did help
quite a bit, I am sure :)
Resolve produced comparable maps that were 'subjectively' slightly less
pretty. As a crude measure of
This is just one option out of many, but what often works miracles in cases
like this is to subtly alter the DNA oligo - add or remove 1-2 nt from the
ends, or to methylate/phosphorylate/dephosphorylate etc. This is quite cheap
for short oligos and therefore can be very cost effective.
Artem
The easiest (albeit by far not the simplest!) option is to mutate the
offending amino acid to Gly, then back to what it should be.
Alternatively you could use a script to accomplish the same, but if you only
messed up 1 or 2 residues, the above is easier.
Artem
-Original Message-
From:
I would assume that the wrong chiralities were introduced during building.
It is not very hard to change chiral centers around if you're dragging atoms
around in and out of density...
Artem
-Original Message-
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Dale
Tronrud
Defining specific restraints on bonds connecting the chromophore with the
rest of the protein worked fine for us in Refmac when we solved a turboGFP
structure a couple of years ago. There wasn’t anything special that had to
be done �C all strictly by the book…
Artem
_
From: CCP4
Hi,
I assume that there is no other way and you have to share the space, so I
will save you from the standard tirade about functional space segregation
:-)
If the air in the lab is so loaded with solvents that your plates are
melting - then *you* (or anyone else for that matter) certainly
Hi Mark,
0.
1. Please note that you don't HAVE to have NdeI/ClaI cuts in your vector, as
both enzymes have several compatible end cuts. It's not as good for NdeI but
ClaI has decent numbers of compatible ends.
2. Unless your gene is super-huge or really difficult to PCR, I see no
reason to avoid
Hi,
(still am confused about how far or how much a commercial people(non
academic) can access this BB)
Most of the people I know in industry at least occasionally read this board.
Not everyone posts of course, but that's nothing to do with your affiliation
- not everyone from academia posts
Tryptic digest of the excised gel band, followed by rigorous peptide
matching helped us identify a fragment in a very recent case. The
crystallographic results confirmed MS findings.
Curiously, the ends were 'ragged' so direct ESI-MS was not very useful,
likewise the N-terminal sequencing would
,
Does high salt concentration(around 0.7M Nacl) in NiNTA elution do better.
And what about the purification of high positively charged protein (like PI
10) in NiNTA.
Thanks
Debajyoti Dutta
On Sat, 28 Jun 2008 Artem Evdokimov wrote :
Ni salt of dodecyl sulphate is not soluble. Therefore (at least
Ni salt of dodecyl sulphate is not soluble. Therefore (at least in theory)
SDS may leach the Ni out of the chelate and deposit it throughout the column
in baby-blue soapy flecks. Having said that, I must add that if you have to
use more than 0.1% SDS then you're dealing with a truly extreme case!
Concentration of large volumes is a common bane of many biochemistry/protein
production labs. Diaflow can be quite expensive and regular concentrators
tend to take forever. Thus, my preferred solution is not to use
concentrators at all.
1. AS cuts do not really *require* high protein
Highly recommended:
Animal Cell Culture: A Practical Approach
by R. Ian Freshney (Editor)
Artem
-Original Message-
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Exec
Sent: Tuesday, June 24, 2008 10:59 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] cell line
Hi
to
dialize against a high concentration of monovalent
salt first, not
just deionized water.
On Jun 22, 2008, at 9:10 AM, E rajakumar wrote:
Hi Artem Evdokimov
Thank you for the mail. I have synthesized DMT-on
oligos in our laboratory. Deprotection was
performed
treating with ammonium
Hi,
How did you synthesize the DNA? I assume external vendor (so few people make
their own these days)? How was the DNA purified? Sometimes if only a
'desalting' step is used there may be 'other chemicals' in the mix. Also,
what pH was your DNA at, and in what buffer (if any)? If your DNA
Mercurochrome (mebromin) is both a derivative AND a stain (tested, true).
It's just like having a pie and eating it at the same time.
Artem
-Original Message-
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of
Richard Gillilan
Sent: Sunday, June 15, 2008 9:36 PM
To:
Concanavalin A can be made to bind Zn and Ca. Its binding site has room for
one transition metal and one alkaline earth metal.
http://www.jbc.org/cgi/content/abstract/271/27/16144
Highly recommended reading material on one of the classical protein targets
Sumner's Nobel Prize lecture is here
Hi,
Others have suggested very rational possibilities, so I would like to
mention the odd one out:
It may sound strange but I have to ask - 50% activity as measured how? Many
biological assays have intrinsic experimental errors around 10%. If you add
on top of this the difficulties encountered
I would instead try for a *shorter* equilibration period, possibly with just
an hour or less. Or even try seeding into batch.
Artem
_
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of amit
sharma
Sent: Saturday, May 31, 2008 5:42 AM
To: CCP4BB@JISCMAIL.AC.UK
Hi,
Unless you've already tried exhaustively - why not to try MIR, as you
mentioned - there are many derivatives out there to be tried and with a
small protein the chances are pretty good. All you need is one good
derivative :)
This certainly sounds easier than mutating exposed Met!
Hello Sajid,
The crosshairs are chip boundaries on a multi-CCD detector. No need to mask
them.
Your lattice appears to be twinned, on cursory inspection. Also you have bad
cryo - I bet if you improve your cryo, the resolution of observable
diffraction increases by at least 0.3A.
Good luck,
Hi,
This is not a single-step procedure and as far as I know there are several
ways to do this - but I would recommend first to build a model of the
domain(s) you're interested in. That is, if you can do that. Based on the
model you should be able to derermine the first (last) amino acids
About 2 - 2.5 minutes on Mosquito for single-drop protocol, scaled according
to the number of drops per well (7 minutes for 3-drop trays). About 25
minutes on a Tecan-derived platform (numbers vary greatly depending on the
particular configuration).
Artem
_
From: CCP4 bulletin
Hi,
Sorry - my comments do not have much to do with the question you asked.
I would like to humbly request that people refrain from sending
multi-megabyte files through a mail-list server. If you have to share a
large file, a polite way to do so is to use a free file hosting server so
that
Hi,
Generally speaking, these days you don't need a special vector to introduce
a C-His (or an N-His, BAP, STREP, STREPII, or any short tag for that
matter), since the sequence of the tag + cleavage site is short enough to be
engineered directly into your PCR amplicon. There are numerous ways
I've used cheap Peltier incubators in the past (when funds were scarse) and
they were just fine - with the caveat that the incubators I used were small
-- basically they were portable beer coolers with a temperature controller
(made by Koolatron). First thing I did was to check the temperature and
If you have poor conservation in this region and the region itself is a
loop, I would perhaps attempt to replace the entire loop with another one -
from a homologue that does not have the glycosylation site(s). Not that this
is guarranteed to work, but it might.
Artem
-Original Message-
There's an even faster reagent for staining - takes abut 1 minute overall
and the results look just like dear old Coomassie. This reagent works
wonderfully for us. Yes, the recipe is proprietary - but if you add up the
cost of reagents for normal Coomassie and compare - this one isn't much more
Dear Eric,
There are several essential variables that govern protein chromatography
(whether affinity, ion exchange, sizing, or other). It would be silly of me
to reproduce a protein chromatography handbook here, so instead I would just
list some practical pointers:
If your column is e.g. 10-15
Jacob,
You can try several things, including the stuff already mentioned by others
- EDTA, salt, etc. A very useful option to keep in mind is to check that
excess imidazole isn't causing the problem. You can find out by simply
diluting the fractions down, or by changing the resin. His-SELECT
Hi,
I would like to point out, for the sake of fairness, that thrombin (high
quality bovine thrombin such as sold by HTI for example) is still much
cheaper *to use* than commercial TEV. One milligram of TEV, TVMV, AcTEV, and
so forth can be used to cleave anywhere in between 10 to 100 mg of
Hi Kendall,
As to (1) - it's a hard task. Redox potentials of Cys residues vary with
their environment, so it is very difficult to predict what might happen in
any specific case. Mixtures of reduced and oxidized GSH are commonly used to
maintain a specific redox environment, however if you're
Yangming,
This topic has been discussed before - basically there are no easily
discernable trends - some proteins crystallize better with a tag, others -
without, and yet others - don't care whether the tag is there or not. I tend
to try either, just to see what works better (time and effort
In my humble opinion starting with a mixed culture is a bad idea, because:
Successful expressors have heightened metabolic load even in the absence of
inducing agents. Therefore, during cultivation their 'expression-less'
companions will progress more rapidly and undergo more cell divisions -
Hi,
For E. coli, as others already pointed out - optimizing codon usage of a
gene 'does not usually hurt'. I can attest to the fact that it can boost
expression quite a bit - at least in the modest number of cases where I have
had a chance to compare the expression levels under the same
Brenda,
Generally speaking it is not abnormal to at least *hope* for co-crystals to
appear at or around the condition(s) where the native crystals are grown.
Therefore many people in fact begin looking for co-crystals by screening a
fine grid around the expected crystallization conditions for the
Or use print ` xxx `
Artem
-Original Message-
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Gerard
DVD Kleywegt
Sent: Monday, January 14, 2008 5:24 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] PERL system call to CCP4
hi,
I am writing a PERL script to execute
This depends of course on your definition of signature motifs, taking this
in a broad sense - many proteins that have multiple functional domains
probably fit the bill, so you could search for really huge proteins.
For example:
FAS (human, not bacterial) - 2504 amino acids in a single chain (!)
Hi,
Heavy atoms can be unpredictable. There are rules of thumb regarding their
use but these rules are frequently bent and broken.
Which heavy atom derivatives have you tried, and at what (approximately)
concentrations and exposure times? Does your protein have residues that
*can* react
You should check if this residue is a part of a glycosylation pattern
(either via crude computational sequence analysis, or if you're lucky - by
digest and peptide MS).
The density *looks like* a disordered sugar but at 2.6A this is a very tough
call.
Artem
_
From: CCP4 bulletin
Phoebe,
It just so happens that we've been working on a C4 Zn-finger (or something
very much like a Zn-finger) very recently without knowing that we had one
(it's in the PheRS from S. haemolyticus, link to paper below) and we've
purified the protein w/o any issues on an IMAC column (His-SELECT).
What's the resolution? At low res it is possible to miss a subtle movement
of e.g. some sidechains which are being replaced by the ligand. What quality
parameters did the MR have? Can you omit the entire ligand binding site,
refine, and re-generate the map - what does it look like after that?
A
This is a tough question. Certainly, in an ideal world (and also in the
world that e.g. Hampton or Emerald salespeople would like us to live in) you
should replace expired reagents. The reality of course is that it's
sometimes quite costly to do so. It is also expensive (in terms of time and
:
Artem Evdokimov mailto:[EMAIL PROTECTED] [EMAIL PROTECTED]
Reply-To:
Artem Evdokimov mailto:[EMAIL PROTECTED] [EMAIL PROTECTED]
To:
CCP4BB@JISCMAIL.AC.UK
References:
mailto:[EMAIL PROTECTED]
[EMAIL PROTECTED]
This is a tough question. Certainly, in an ideal world (and also
Dear Jerry,
One way I can think of would be to try the magical polymer NV-10 sold by
Novexin (UK). It does actually work very well for proteins with low
solubility. You'd add a few mg/ml of polymer to each protein solution, mix
the two and see what happens. This polymer has already saved
If you send a picture of this stuff to the list, I bet someone may recognize
it :)
I think it's called 'distributed computing'.
On a more serious note, a pharmacophore matching routine would do what you
need, but you'd have to convert from E.D. to a 'blank' pharmacophore with
all property
Vijay,
It is hard to guess what you mean when you say that 'mass spec results
confirmed both protein bands are the same'. Do you mean that both bands were
cut, digested with e.g. trypsin, and the resulting peptides were mapped? Or,
alternatively, you've done an MS spectrum on the sample that
Yes!
And if you have money, you can use phosphoramidon
http://delphi.phys.univ-tours.fr/Prolysis/phosphoramidon.html
It's nearly as good as EDTA or phenantroline for inhibiting metalloproteases
- and it's fully compatible with IMAC!
Artem
-Original Message-
From: CCP4 bulletin board
Is there an option to include a small inset in the corner with a structure
diagram? CPK colors do not always work well when obscured by e.d. mesh.
Artem
-Original Message-
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of
Yanming Zhang
Sent: Monday, October 29, 2007 1:36
I don't like using 'carve' options but I can see where it may come useful.
On the positive side, in the age where deposition of coordinates and
structure factors is almost mandatory, there shouldn't be a question of how
your map looks because presumably the skeptical reader can go and calculate
Cannot beat the price of a TriTek imager - CrystalPro. For the price this is
probably the optimum product.
Artem
Note - I am not associated with TriTek and in fact our lab uses Formulatrix
imagers, but these are much more expensive.
_
From: CCP4 bulletin board
Another option would be an Art Robbins imager - also a value-priced
instrument.
Artem
_
From: Artem Evdokimov [mailto:[EMAIL PROTECTED]
Sent: Friday, October 26, 2007 7:08 PM
To: 'Page, Rebecca'; 'CCP4BB@JISCMAIL.AC.UK'
Subject: RE: [ccp4bb] Cost-effective imaging systems
Dear Colleagues,
We would like extend an invitation to you for the 3rd annual Ohio Valley
Crystallography and Biophysics Symposium (now with more biophysics!), to be
held this year at the University of Toledo, Ohio on Friday, November 9th. In
addition to talks on macromolecular structure
Dear Colleagues,
It is with sorrow I report the passing of Gerard (Gerry) Bunick on September
19, 2007 in Oak Ridge, Tennessee after an 11 year struggle with cancer. In
addition to other significant research, Gerry was one of the architects of
the resurgence in neutron protein crystallography.
The presence of a His-tag in the sequence does not automatically mean that
the protein will bind to IMAC resin. Since your protein is so huge and since
the His-MBP-fusion does not work any better than the His-fusion I would
hazard that your protein is highly aggregated in solution, to the extent
If you want to be 'by the book' on it, you should follow IUPAC rules for
atomic numbering of chemical compounds. Unless you're an organic chemist,
these rules are pretty hard to follow, so most people number at will.
If you're dealing with a 'known' series of compounds, such as analogous
By the way, I would like to mention that I was sloppy with the language in
the original reply - I was referring to complex half-life (T1/2) which is
expressed in direct units of time, not the kon or koff, which are expressed
in inverse units of time. Thank you Raji Edayathumangalam for pointing
Hello,
While I am not a professional MS person, I have had the privilege of working
with many MS scientists and some knowledge has rubbed off despite my best
efforts to stay benighted. All the correct information in the post below is
therefore credited to the MS community whereas all the
The literature already contains quite a few papers discussing ligand-protein
interactions derived from low-resolution data, noisy data, etc. It's
relatively easy to take a low-quality map; dock the molecule willy-nilly
into the poorly defined 'blobule' of density, and derive spectacular
Hi,
They're likely both BME adducts, just in the first case the CH2CH2OH portion
is way more disordered.
Artem
_
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of
[EMAIL PROTECTED]
Sent: Monday, August 13, 2007 9:59 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb]
Hi,
I can turn it back on as easily as it was turned off - as I said, several
people had negative comments, so I decided to to turn it off for now. If
there's more positive than negative impact - I would be glad to turn it back
on.
Artem
_
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