Do we deposit these anywhere?
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Hello my fellow structural biologists, I am contemplating why some choose
the HRV3C protease site over TEV for their fusion proteins. Does anyone
know? Can HRV3C be made easily in homelab? Does anyone have a plasmid?
Thank you, G
Hello friends, Can anyone recommend an antibody that works well
forbbonding native MBP tag? Many thanks, G
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What is the best way to display Harker sections... these days?
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What is the best way to display Harker sections these days?
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I was thinking the form of GFP that dimerizes. This would also make it
easy to track where the protein is.
On Tue, Sep 22, 2020 at 1:28 PM Diana Tomchick <
diana.tomch...@utsouthwestern.edu> wrote:
> Any dimeric tag should work if you add a long enough linker to satisfy
> your distance
Another protein purification question: Does anyone know what is AcTEV
protease that is sold by Thermofisher Scientific? Is this the same as
SuperTEV?
It works well but is so expensive.
Thanks for any advice, Gloria
To
We were looking at these, they look like fun.
https://www.wetkeys.com/Soft-touch-Comfort-Hygienic-Washable-Keyboard-USB-p/kbstfc106-w.htm
On Tue, May 5, 2020, 7:20 PM James Holton wrote:
> All joking aside, there has been a furor of attention on UV-based
> disinfection of late. Some of it
Hi Friends, We are secreting Spike Ecto domain into the media from insect
cells for purification. As we scale up I am wondering what is recommended
for collecting the media from large volumes of culture. Centrifugation?
Filtration of some kind? I imagine we need to be gentle to not lyse the
I personally don't tweet.
On Tue, Mar 31, 2020 at 12:21 PM Sweet, Robert <
27e0eb9d20ec-dmarc-requ...@jiscmail.ac.uk> wrote:
> Real Men (and possibly Women too) Don't Tweet.
>
> Bob
>
>
> From: CCP4 bulletin board on behalf of James
> Holton
>
Actually we don't declare anything to TSA, Jahaun just carries them in his
carryon and it is fine. Easy peasy?
On Wed, Feb 19, 2020 at 3:48 PM <
0c2488af9525-dmarc-requ...@jiscmail.ac.uk> wrote:
>
> ... and don't say you're travelling with heavy water!
>
> Jon Cooper
>
> On 19 Feb 2020
Hello CCP4-ers,
I was wondering what people have found to be the best human cell line
expression system for making a large quantity of purified recombinant
protein.
Any information and protocols would be greatly appreciated.
Happy 2020, Gloria
mistry, and Medicine
(2) Cell Signaling at Membranes
(3) Macromolecular Complex Assembly and Disassembly
(4) Bioinspired Materials and Technology
We are looking forward to your participation!
On behalf of Biology Session Chairs:
Dr. Gloria Borgstahl
University of Nebraska Medical Center
Dr. Y
Dear friends in crystallography,
I know this seems unrelated, but it really isn't ... please forgive me.
We are trying to use the Avitag/BirA system to specifically biotinylate
target proteins in an economical manner. Are any of you purifying the BirA
enzyme in your lab for biotinylation and if
Sorry to be late chiming in on this post (survived RAGBRAI). I think the
challenges (crystallization, perdeuteration) and benefits of neutron
crystallography (where are those protons) could be included. We are now in
an era of using cryotrapping with neutrons which I think is really cutting
edge
You might want to quote how many researchers use the PDB instead. Maybe an
inquiry to the PDB could give you that statistic.
That would give you a big number
On Wed, May 29, 2019 at 1:54 PM Scott Horowitz wrote:
> Hi all, I was recently asked how many biological
> crystallographers plus
Hi Matthew, I am also a bit late in responding, I have a few
incommensurately modulated protein crystal datasets that you would be
welcome to use in your course. It would be neat for students to at
least know that this type of diffraction exists. As far as I know,
they can only be processed
Hello, friends in crystallography,
A colleague just asked me this question. He is worried about trace
protease interfering with the receptors he is studying in cell-based
experiments using a 110 amino acid protein we made for him. He has
been unable to make the peptide synthetically. The
I am very interested in this topic, and have found incommensurately
modulated crystals and few examples of possible protein quasicrystals.
Do you have any images you could share?
On Tue, Feb 13, 2018 at 9:09 AM, Yu Qiu wrote:
> Hi,
>
>
>
> I have been trying to crystallize a
My answer is not precise, I just try to give students something they
can remember, so they don't go and model water for example at an
inappropriate resolution. Of course phasing matters as well.
On Thu, Jan 11, 2018 at 2:31 PM, Keller, Jacob wrote:
>>I tell people it
Doe any one know what resolution is needed to distinguish phosphate
from carbon in a difference map?
I tell people it is when your resolution is less than the bond length
that connects the two atoms.
On Thu, Jan 11, 2018 at 1:59 PM, Thomas Edwards wrote:
> Dear Jacob,
>
> Ah... this old chestnut!
>
> Current EM people say that they are at atomic resolution because they
I have seen this in MnSOD. We had a tetramer in the ASU and each
active site had different ligands in our peroxide soak. I assumed the
crystal lattice can influence these things or it is inappropriate
metal incorporation. I would highly recommend doing ICP-MS on a
dissolved crystal and on your
If you mean you truncated the low resolution data to 5 angstrom, I
wouldn't recommend that.
On Fri, Nov 3, 2017 at 12:24 PM, Eleanor Dodson
wrote:
> That map does not look like a 5A map?
> I guess you mean something else..
> Eleanor
>
> On 3 November 2017 at 11:00,
Our RPA14/32 crystals from 2007 included full length protein for both
subunits (RPA14 and RPA32), but only the central OB fold of RPA32
could be modelled. The crystals included RPA32(1-270) but only 42-176
could be modelled. I remember being very frustrated by not being able
to visualize the
Thanks James, Yes Charles I have an active NSF grant on this topic.
We should talk, Gloria
On Mon, Sep 18, 2017 at 11:05 AM, James Holton
wrote:
>
> I believe Gloria Borgstahl's lab at UNMC has done a little bit of work on
> this.
>
>
> -James Holton
>
> MAD Scientist
--
From: Gloria Borgstahl <gborgst...@gmail.com>
Date: Tue, Jul 11, 2017 at 1:04 PM
Subject: Granada Crystallization Boxes?
To: "CCP4BB@JISCMAIL.AC.UK" <CCP4BB@jiscmail.ac.uk>
I have recently found out that these are no longer being manufactured
or sold commercially. But,
ne might
> have some.
>
> Best wishes, Patrick
>
>
> On 11 July 2017 at 19:04, Gloria Borgstahl <gborgst...@gmail.com> wrote:
>
>> I have recently found out that these are no longer being manufactured or
>> sold commercially. But, as fortune has it, we have ju
gt; Rm. ND10.214A
> Dallas, TX 75390-8816
> diana.tomch...@utsouthwestern.edu
> (214) 645-6383 (phone)
> (214) 645-6353 (fax)
>
> On Jul 11, 2017, at 1:04 PM, Gloria Borgstahl <gborgst...@gmail.com>
> wrote:
>
> I have recently found out that these are no longer being manuf
I have recently found out that these are no longer being manufactured or
sold commercially. But, as fortune has it, we have just been funded to fly
some large quartz capillaries crystallization experimente up to the
International Space Station for neutron crystallography. Our experimental
design
, etc. If you can, please register to present a research poster. The
associated journal club is included in the website and will be broadcasted
remotely if you wish to participate from your office.
Thank you for your interest, Gloria Borgstahl and Luis Marky
Please share this email.
To register
I would recommend just mounting them in capillaries without loops, as this
will travel better.
Put slugs of mother liquor on either side, seal with wax and then coat wax
and onto the end of capillary glass lightly with fingernail polish.
We have had terrible luck with FedEx delivering damaged
I have learned to refrain from catapulting my crystals.
Happy holidays, God bless us ... everyone (with diffracting crystals)
On Wed, Dec 21, 2016 at 2:35 PM, Keller, Jacob
wrote:
> >In a second case we were working on a beamline on the west coast of the
> US. Every
Eddie Snell for President
On Wed, Nov 9, 2016 at 11:02 AM, Edward Snell
wrote:
> As a Brexit and Trumpet affected person having a foot in both countries
> ,this topic is too far off the normal discussion on CCP4 and probably
> better taken up privately. CCP4 is not a
Did they stop supporting it due to lack of renewed funding and having to
cut staff that had the knowledge?
I'm pretty sure you only know part of the story.
On Tue, May 12, 2015 at 11:48 AM, James Stroud xtald...@gmail.com wrote:
I hereby call on the broadest community of academics and
High concentration of ammonium formate is a cryosolvent
On Mon, May 4, 2015 at 5:20 PM, Tristan Croll tristan.cr...@qut.edu.au
wrote:
What about nature's favourite cryoprotectant, trehalose?
From: CCP4 bulletin board CCP4BB@JISCMAIL.AC.UK on behalf of
Ah Ha!
drum roll
A quasicrystal !
On Wed, Apr 1, 2015 at 7:00 AM, Julia Griese gri...@dbb.su.se wrote:
This one appears to be of a similar age. It has a most puzzling, but
pretty pentagonal pattern (and a backstop). Unfortunately Mosflm doesn't
appear to support the image format.
I have had young ones grow lysozyme crystals in just a few minutes, using
eye droppers and petri dishes. The crystals grow very fast, you can watch
them grow in the microscope. Also they grow large enough you can see them
by eye. Some izit dye would be fun to add (never did that). Then I let
Apparently these are not for sale anymore by EMD Millipore
yet it is what is recommended for plasmid preps with pETcoco-2.
Does anyone know of a source of these cells
or a substitute cell line we could use?
Thanks, Gloria
Thanks Mark, this is a good tip
what would you use for Rosetta cells though?
Tetracyclin?
On Fri, Oct 17, 2014 at 8:31 AM, Mark Wilson mwilso...@unl.edu wrote:
Hi Ivan,
We've had good luck with the addition of chloramphenicol ~1 hour prior to
harvest, as described in:
Carrió, M. M., and
Be aware that some buffers are temperature sensitive and change pH, if this
pH change heads toward the pI of the protein it can crash out.
On Tue, Aug 19, 2014 at 6:37 PM, Prince, D Bryan
dbryan.pri...@astrazeneca.com wrote:
Dear Prashant,
I have been working with a protein-protein
Does any one know of a source of these cuvettes?
Protein Solution doesn't exist anymore
and Wyatt no longer has these.
You can prevent them from falling off by also
removing 5 microliter or so of mother liquor from the drop and
repositioning it back over the reservoir
On Wed, Jul 2, 2014 at 3:38 PM, Patrick Loll pat.l...@drexel.edu wrote:
You can cut a small piece of sponge and put that into the reservoir;
I vote for Z's idea
On Wed, May 14, 2014 at 12:32 PM, Zachary Wood z...@bmb.uga.edu wrote:
Hello All,
Instead of placing the additional burden of policing on the good people at
the PDB, perhaps the entry page for each structure could contain a comments
section. Then the community could
Made my day
Her biography was inspirational to me!
On Mon, May 12, 2014 at 9:52 AM, lbetts0508 . laurie.betts0...@gmail.comwrote:
I should have said, ALL crystallographers rejoice!! Laurie
On Mon, May 12, 2014 at 10:01 AM, lbetts0508 . laurie.betts0...@gmail.com
wrote:
Oh
You can increase the formate concentration and it will be a cryoprotectant
on its own. Test increasing amounts until you find the concentrationthat
freezed clear as glass and the transfer your crystal to this. We did this
with sodium formate at 7 M quite easily.
On Tue, Dec 17, 2013 at 2:59
specific. If you don't get a better lead I can send you the
sequence and a reference.
Gloria Borgstahl wrote:
Does anyone know of a good way to phosphorylate the Tyr on a protein for
structural
studies? Is there a generic kinase that can be coexpressed or purified
for phosphorylation
Does anyone know of a good way to phosphorylate the Tyr on a protein for
structural studies? Is there a generic kinase that can be coexpressed or
purified for phosphorylation?
The pCMF Amber codon system is very expensive
and Glu really doesn't mimic pTyr all that well.
Any ideas/help would be
We had this happen with RPA14/32 heterodimer, we kept the two peaks
separate, and then grew crystals from each but in different space groups
and diffraction resolution. See Habel, J. E., Ohren, J. F. and Borgstahl,
G. E. O. Dynamic light scattering analysis of full-length, human RPA14/32
dimer:
Are you talking about the Fig 3.1 in their book, Laboratory-build
oscillation camera (Arndt, Champness, Phizackerley and Wonacott, 1973)
On Thu, Oct 31, 2013 at 2:28 AM, Eleanor Dodson
eleanor.dod...@york.ac.ukwrote:
one in their book, i am sure.
eleanor
On 30 Oct 2013, at 16:05, Gerard
We are trying to apply this approach for the first time to define a
crystallizable domain of our protein of interest. Can anyone send a
protocol that includes exactly how to do the mass spectrometry
measurement? Our core lab here doesn't know and I'm just not that gifted.
Happy October, G
Hello ccp4ers,
I am helping a colleague develop a grant and have a vague recollection of
structures of transmembrane protein receptors that signal across the
membrane. Can anyone send me specific examples?
Many thanks, G
We have a protein sequence that probably contains OB folds. What is the
best way to search for the top structural homologs to this sequence in the
pdb? G
We have found funding for a 96 well DLS from Wyatt. I was wondering what
peoples experiences with this instrument was. Is there a better
instrument out there? Does it matter where you buy the 96 well plates from?
Many thanks, Gloria
In our lab we do DLS on the SEC fractions and only set up those that are
monodisperse :-)
On Mon, Jul 1, 2013 at 7:37 PM, El Arnaout, Toufic
elarnao...@biochem.wustl.edu wrote:
Hello Peter,
In addition to the great comments/details, please check the following
points I have now in mind..
We have had good luck with making a protein soluble using the Artic Express
bacterial cell line BUT we can't get rid of the chaperone that copurifies.
We have tried adding ATP, MgCl2 and potassium to lysate and extensive
washes and this releases a bit of the chaperone. Has anyone solved this
I have found it is best to test the absorption of your buffer in the
wavelength range you are interested in.
If you are going to do a temperature study with CD perhaps at 222 nm, then
test your buffer there with your UV spec.
You want to have little or no absorption. Or do the range 200-270 nm
Daniel Schectman's 2012 nobel prize for the discovery of quasicrystals is
missing.
On Sat, Oct 13, 2012 at 5:01 PM, Joel Sussman
joel.suss...@weizmann.ac.ilwrote:
Just want to make sure anyone interested in Nobel Prizes knows about this
existing page:
This indeed is sad news for today.
I just wanted to note that Professor Johnson's early papers on
time-resolved crystallography truly inspired me to continue in
crystallography, influenced my decision for my first postdoctoral position
and to push the limits. I still have the carefully
How would I tell REFMAC to replace the
C-terminal Pro in my synthetic peptide
with a CONH2
not
the natural CT.
--
From: Gloria Borgstahl gborgst...@gmail.com
Date: Thu, Aug 23, 2012 at 12:25 PM
Subject: CONH2 not COOH at TER
To: ccp4bb@jiscmail.ac.uk
How would I tell REFMAC to replace the
C-terminal Pro in my synthetic peptide
with a CONH2
not
the natural CT.
We used VOIDOO recently and it worked well
Kleywegt, G. J. Jones, T. A. (1994). Detection, delineation,
measurement and display of cavities in macromolecular structures. Acta
Crystallogr. D Biol. Crystallogr. 50, 178-185.
On Thu, May 31, 2012 at 12:41 PM, Zhou, Tongqing (NIH/VRC) [E]
My fellow crystallographers,
We are thinking the SUMO/His vectors would be nice to have in the lab
aresenal... but. The stumbling block is that the protease needed for
cleavage is very expensive at crystallography scale. SUMO(ULP-1)
protease costs ~$700/mg fusion protein. It would not be a
a recent experience in our lab with molecular replacement (wt and
disordered point mutant; same space group and unit cell)
was solved with a combination of two methods.
1. We made omit maps in the disordered region at several lower
resolutions. The region became interpretable after suffereing
Hello all,
We are solving a superstructure of a protein complex with 2 parts.
Built 6 of the first part and they are all sensibly stacked next to each other.
Then we read this into molrep as the fixed model and solved for the
second part.
The solution was found but the 6 for the second model are
A thing we frequently forget is that phosphate can be a precipitating agent
try a phosphate grid screen, just like you would with ammonium sulfate.
If your protein likes PBS, it may want to crystallize with phosphate
See Enrico Stura's footprint screen for example
On Tue, Nov 15, 2011 at 5:25
Glutaraldehyde works best at low pH
On Mon, Nov 7, 2011 at 8:40 AM, Ed Pozharski epozh...@umaryland.edu wrote:
On Mon, 2011-11-07 at 05:19 +, Sam Arnosti wrote:
Hi everyone
I have a protein that is extraordinarily stable at PH=3.0 or even 2.0.
I want to crystallize it in the low PH and
I just want to jump in to state that I am ALL FOR the notion of
depositing the images that go with the structure factors and the
refined structure.
Through the years, I have been interviewing folks about the strange
satellite diffraction they saw, but ignored,
used the mains that they could
Dale, This is exactly the conversation I just had with my student Jason,
right on!
The paper we are writing just now, this is figure 1.
But I always get rejected by Nature, so go figure.
On Thu, Aug 11, 2011 at 1:25 PM, Dale Tronrud det...@uoxray.uoregon.eduwrote:
I agree with Prof. Tomchick:
Hi Guys,
we are learning to work with Sf9 cells and Carol in my lab wanted me to ask
you the following question. Many thanks for any help, G
I need to titer a baculovirus stock in my suspension-adapted Sf9 cells. I
know that these can be encouraged to attach better to tissue culture plastic
if
I'm voting with Roger this time.
If I were you I would model a nickel in there (unless you have a better
candidate)
if it is right, the distances to the His should be like seen in a SOD active
site.
Then you can model the bonds and waters. You may need partial occupancy on
the metal.
Reminds me
We are using TEV protease to make a big batch of protein for
structural studies. We usually use thrombin, so this is our first
time using this enzyme on a large scale. Boy is it expensive! Does
anyone know of a bulk source for this enzyme and what ratios of use do
you recommend. Imidazole
I have learned alot about TEV protease
and thank you all for your help
Going to start making it ourselves!!!
of a disulfide bond on a
2-fold connecting two homodimers).
So I polled the collective knowledge of the great ccp4bb group.
On Wed, Dec 8, 2010 at 10:57 AM, Gloria Borgstahl gborgst...@gmail.com wrote:
My fellow crystallographers,
I wanted to take a poll.
How many of you have ever had a protein
.
**
Gloria Borgstahl
Eppley Institute for Cancer Research and Allied Diseases
987696 Nebraska Medical Center
10732A Lied Transplant Center
Omaha, NE 68198-7696
http://sbl.unmc.edu
Office (402) 559-8578
FAX (402) 559-3739
Here was the query.What is the easiest way, these days, to calculate the buried surface area between two subunits of a protein? Here are the software and links received (thank you all):1.4+ votes for PISA - This following website is great and does a very nice job of analyzing protein interfaces
Hello all,
What is the easiest way, these days, to calculate the buried surface area
between two subunits of a protein?
Thanks, and Happy Friday,
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