For protein-DNA complexes, we generally found that the ends of the DNA duplex
are the most important variable because DNA bases love to stack and usually
form crystal-packing contacts. Instead of more screening kits (say, one
fairly low salt PEG-ion screen kit should do) buy lots of oligos tha
For plasmids, I think the best long-term option is to send them to Addgene (as
long as they’re ready for public use).
From: CCP4 bulletin board on behalf of Hughes, Jonathan
Date: Wednesday, October 9, 2024 at 2:38 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] AW: [ccp4bb] Commercial storage
departments, one on each plan. Don’t assume logic.
~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
https://voices.uchicago.edu/phoebericelab/
From: CCP4 bulletin board on behalf of Hekstra, Doeke
Romke
Date: Sunday, October 15, 2023
-CH3)
~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
https://voices.uchicago.edu/phoebericelab/
From: CCP4 bulletin board on behalf of Eleanor Dodson
<176a9d5ebad7-dmarc-requ...@jiscmail.ac.uk>
Date: Monday, September 18,
Excellent idea.
From: CCP4 bulletin board on behalf of Debanu Das
Date: Saturday, December 24, 2022 at 5:57 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Future Diffraction Methods
Consider merging with the Annual ACA meeting. The ACA meeting can also benefit
from more X-ray diffraction
[Text Description automatically generated]
Postdoctoral Position in the Rice Lab at the University of Chicago.
Mechanism, Engineering, and Structural Biology of Large Serine Integrases
A postdoctoral position is available immediately the laboratory of Dr. Phoebe
Rice.
Large serine integrases (
You might try this:
https://seq2fun.dcmb.med.umich.edu/DeepFoldRNA/
I’m recommending it with the caveat that I’ve tested it on exactly 1
unpublished structure, and it did a really impressive but far from perfect job.
From: CCP4 bulletin board on behalf of Abhilasha Thakur
Date: Thursday, Se
Apologies for the stupid post about freezing conditions last night – my brain
was clearly very low on glucose when I read that list of temperatures!
From: CCP4 bulletin board on behalf of Phoebe A. Rice
Date: Wednesday, September 7, 2022 at 7:49 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re
I guess the big question is what is the question that you’re trying to address
from those numbers? I’d be nervous about making conclusions about trends in B
factors from just 1 data set per temperature. As you probably know, the B
factors will reflect static differences in atomic position acr
Why why why doesn’t REFMAC just write a new column – something like FOBS_mod?
Shouldn’t it be a general rule that if you change something, you give it a
different name?
From: CCP4 bulletin board on behalf of Randy John Read
Date: Wednesday, July 13, 2022 at 5:22 AM
To: CCP4BB@JISCMAIL.AC.UK
Oo that looks handy!
And it reminds me to ask the community: if you wanted to do outreach to high
schoolers and get them to, say, look at a DNA structure, is there a PyMol-like
program that works on tablets? I see PyMol for ipad has been discontinued.
Thanks,
Phoebe
From: CCP4 bulletin boa
I would try the tools on this site - http://web.x3dna.org/index.php/protein
From: CCP4 bulletin board on behalf of "Srivastava,
Dhiraj"
Reply-To: "Srivastava, Dhiraj"
Date: Monday, December 27, 2021 at 8:32 PM
To: "CCP4BB@JISCMAIL.AC.UK"
Subject: [ccp4bb] protein DNA complex structure and ex
A side point, worth considering depending on your x-ray source: many years ago
we threw out several crystals in a row for having split spots before we finally
realized that the BEAM was split, not the crystals. Oops.
~~~
Phoebe A. Rice
Dept. of Biochem & Mol.
Worth discussion although may be impossible to fix.
My 2 cents:
Standard contracts here seem to be for 1 year, but I've always worked on the
assumption that the contract will be renewed unless post-doc and PI have
"irreconcilable differences" or there is an unforeseen funding shortfall.
Then aga
Dear Silvia,
While high Bs can be a sign that parts of your model are incorrect (and
therefore some careful manual rebuilding may be required), for things like
poorly-ordered surface loops, they can simply be truth.
A point worth remembering for newbies: your goal is to produce the model that
Ah yes, fond memories of that mirrored contraption!
There were also wooden contraptions to strap on one's head with image-merging
mirrors in them. We had fun lining up equal-height friends and merging them.
But both of those did have the downside that getting engrossed in electron
density whi
Each PDB entry now has a DOI which should make referencing that structure
easier in your own publication.
From: CCP4 bulletin board on behalf of Firdous Tarique
Reply-To: Firdous Tarique
Date: Wednesday, September 9, 2020 at 1:35 AM
To: "CCP4BB@JISCMAIL.AC.UK"
Subject: Re: [ccp4bb] taking in
Biochemistry and Biophysics Faculty at The University of Chicago
The Department of Biochemistry and Molecular Biology at the University of
Chicago invites
applications for a faculty position, preferably at the assistant professor
rank. We seek scientists
studying macromolecular dynamics and st
This advance brings deep new meaning to crystallographic data having both a
real and an imaginary component!
On 3/31/20, 11:36 PM, "CCP4 bulletin board on behalf of Petr Kolenko"
wrote:
Dear colleagues,
We, the developers of a program for paired refinement, have found a
remarkable fe
While we're at it, it would also be a great improvement if the PDB would list
nominal resolution in all 3 directions for structures with significantly
anisotropic data. I'm never 100% sure what to do in those cases.
Thanks,
Phoebe
~~~~~~~
Phoebe A. Ric
overstated resolution.
~~~~~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
https://voices.uchicago.edu/phoebericelab/
On 2/28/20, 6:56 AM, "CCP4 bulletin board on behalf of Malý Martin"
wrote:
Dear colleagues,
two different surfaces of B, such that some number (< or = 6) of As could
bridge two B hexamers … or the other way around … Mother Nature is inventive –
best to get some experimental numbers!
~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Commi
While there is some truth to that argument, the problem is that it is harder to
achieve an international reputation in the first place while being routinely
overlooked.
~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
h
With a little fiddling and patience, one can just point a cell phone camera
down the eyepiece. It doesn’t produce the best pics, but works fine if you
just want to stick a few in a notebook (or a grant …).
From: CCP4 bulletin board on behalf of Sergei Strelkov
Reply-To: Sergei Strelkov
Date
function and shows nice
packing.
~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
https://voices.uchicago.edu/phoebericelab/
From: CCP4 bulletin board on behalf of Robert S
Phillips
Reply-To: Robert S Phillips
Date: Wednesday, Nove
cooperative interactions among multiple flexible partners.
~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
https://voices.uchicago.edu/phoebericelab/
From: CCP4 bulletin board on behalf of John R Helliwell
Reply-To: John R Helliwell
a clear, concise log of what kind(s) of density modification were applied?
I get frustrated when other people's students can't tell me exactly what
they've already tried, but it isn't always their fault.
Best,
Phoebe
~~~
Phoebe A. Rice
Dept. o
a...@uchicago.edu> with
their request.
~~~~~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
https://voices.uchicago.edu/phoebericelab/
To unsubscribe from th
At least last fall, the state of the art for getting your T7-transcribed RNA to
start with a triphosphate in phenix was to add this file (edited for your
chains, of course) through the gui (if you use the gui).
Thanks to Deepak Koirala and Nigel Moriarty for help in working this out.
* Phoeb
Very good point, and a good argument against the current trend of publications
in the glossies including everything from mice to omics to structure all in one
manuscript with one set of authors. Especially since it is being pointed out
more vociferously these days (as it should be) that accepti
er as
a buffet of (often quite useful) suggestions for how to rebuild your refmac- or
phenix- refined model.
~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
https://voices.uchicago.edu/phoebericelab/
From: CCP4 bulletin board on beha
-1-Reptangles/dp/B00392NSQ4/ref=sr_1_2?ie=UTF8&qid=1549988668&sr=8-2&keywords=reptangles
You can assemble an amazing array of symmetries with these things.
~~~~~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
https://voi
.
If you're desperate, and your weak spots are weak enough, it is worth a try!
~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
https://voices.uchicago.edu/phoebericelab/
On 1/10/19, 12:18 PM, "CCP4 bulletin board on beh
(with
exceptions of course).
If you only have an existing structure for, say, half of what's in your
asymmetric unit, SAD phases will be less biased than model phases from
molecular replacement, even though both may be noisy.
~~~~~~~
Phoebe A. Rice
Dept. of Biochem &
already come up with a good
solution – either an easy way to install pymol, or an alternative
undergraduate-user-friendly, free macromolecular graphic program that will work
for chromebook users.
Thanks,
Phoebe
~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol.
Sadly Rafal is right the unit cell dimensions don't make sense - either the
space group is wrong or the contents have been digested before crystallization.
The size of the overall unit cell is ~21 x 23 x 43. Given a (DNA-centric but
close enough) view that a duplex is ~25A wide and there are 3.4
How do your twinning statistics look? It could be that the real space group
has lower symmetry (and thus more space for your G4 DNA).
~~~
Phoebe A. Rice
Dept. of Biochem & Mol. Biol. and
Committee on Microbiology
https://voices.uchicago.edu/phoebericelab/
In addition to the other useful advice offered, I’d also suggest determining if
the whole complex or just the DNA fits in the asymmetric unit. Particularly if
the crystals are hexagonal rods or plates, you might have DNA-only crystals.
~~~
Phoebe A. Rice
Dept. of
I'm fond of RaptorX
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
pr...@uchicago.edu<mailto:pr...@uchicago.edu>
https://voices.uchicago.edu/phoebericelab/
From: CCP4
ne is in 3 different rotational states even if there's rough density for
the planes of the bases?
++++++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
pr...@uchicago.edu
https://voices.uchicag
Even though the protein should be denatured by all that urea, we find such gels
sometimes look nicer if stop the reaction with SDS and protease K, and/or
phenol extract the remains of the protein before loading the high-urea gel.
++
Phoebe A. Rice
Dept
You might also be getting aggregation.
If you do an old-fashioned EMSA ("gel shift") assay, does it hang up in the
well?
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
pr...@uchicago.edu<mailto:p
ter.
++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
pr...@uchicago.edu<mailto:pr...@uchicago.edu>
Mobile DNA III: mobile elements are everywhere!
http://www.asmscience.org/content/book/10.1128/
Sorry if this is an insulting question, but did you store it in enough glycerol
to prevent it from freezing? Proteins don't like freeze/thaw cycles,
especially slow ones.
From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of jai mohan
[0cab66323371
This is not at all conclusive, but one depressing thing is that DNA-only
crystals are often hexagonal (like yours are).
Can you reproduce them without the protein?
++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicag
In the distant past, we did get some additional phasing signal by a simplified
version of the RIP concept based on the lability of the bromines in a
5-bromo-dU substituted DNA in an x-ray beam.
We collected the peak anomalous signal first and fast, and then just kept
collecting (we may probably
Hi,
Sometimes automated model building needs more manual intervention than one
might expect, although it sounds like you've already carefully inspected the
"good" regions.
Could you use a model of the N-ter (from a homolog) simply to create a
solvent envelope, then see if solvent flattening
We had a complex many years ago that could sometimes be convinced to
crystallize by poking the drops (probably with a hair): if I remember
correctly, both poking some of the skin into the drop and pulling some of the
oily goo into more contact with the aqueous phase. Then again, that complex
a
At long last, a brilliant theoretical framework for observations previously
dismissed as mere discrepancies!
This being Saturday, can the postulated time-dependent decay of previously
observed observables also be applied to explain the apparent discrepancy in
number of socks entering the wash vs
Thank you Bernard.
I'd like to add that carving the density shown in a figure (especially without
explicitly describing the carve radius used) is the moral equivalent of showing
only 1mm of background above and below the band of interest in a western blot.
Phoebe
_
For historical perspective:
Molecular structure determination by electron microscopy of unstained
crystalline specimens
PNT Unwin, R Henderson - Journal of molecular biology, 1975
http://www.sciencedirect.com/science/article/pii/0022283675902120
From: C
Are you sure there really are 4 chains? 50% solvent may be average, but we've
had crystals with closer to 80%.
From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Pooja Kesari
[pkesar...@gmail.com]
Sent: Tuesday, January 24, 2017 10:42 PM
To: CCP4BB@
Excellent point. I've heard apocryphal stories of someone obtaining beautiful
salt crystals (magnesium ammonium phosphate) by trying to crystallize an ATPase
with ADP and Mg++ using (NH4)2SO4 as a precipitant. Wait long enough, and the
ADP becomes ATP plus AMP, the ATPase takes the 3rd phospha
I found that one can get RSRZ to go way down by loosening the geometry
restraints. The result is a crappy structure and I don't recommend doing that,
but it does get all the atoms crammed into some sort of density.
RSRZ, in my most humble of opinions, seems like one of those statistics that is
Back in my grad school days, we had crystals (of gamma delta resolvase large
fragment) like that: at high protein concentrations, we'd get P6422 with
1/asymmetric unit, and at lower concentrations, C2221 with 3/asymmetric unit
(with an imperfect ncs 2fold 60 degrees from the xtal fold, and very
Interesting way to look at it. But those loop residues are really in the
crystal with an occupancy of 1, so wouldn't letting the B factor fly give a
clearer description of what's in the crystal? Especially as many people know
to color the structure by B factors to get a feel for which bits are
ride during purification)?
Good luck,
Phoebe Rice
++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
pr...@uchicago.edu<mailto:pr...@uchicago.edu>
For one of our more easily-degraded proteins, we added a mix of protease
inhibitors (those expensive tablets you can buy) and also put a drop of EDTA
into each tube in the fraction collector so that any contaminating
metal-dependent proteases would be knocked out as soon as the protein came off
ructure is problematic, please do let the community know!
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
pr...@uchicago.edu<mailto:pr...@uchicago.edu>
From: CCP4 bu
t. If
you can't see nice density for side chains then you probably aren't really
seeing density for waters either.
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
pr...@uchicago.edu&l
ering those of you who are already more up-to-date!
++++++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
pr...@uchicago.edu<mailto:pr...@uchicago.edu>
From: CCP4 bulletin boar
When trying to adjust the "chi" angles of a dT in coot 0.8.1, the methyl and
its H's (which I called C5M to make phenix happy) rotate with the sugar,
producing a rather base bizarre geometry.
nd).
Good luck!
++++++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
pr...@uchicago.edu<mailto:pr...@uchicago.edu>
From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Sudipta
Bhattacharyya [sudiptabhatt
iant pseudo translation."
Phoebe
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu<mailto:pr...@uchicago.edu>
http://bmb.bsd.uchicago.edu/Faculty_and_Research/
ht
print.
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu
http://bmb.bsd.uchicago.edu/Faculty_and_Research/
http://www.rsc.org/shop/books/2008/9780854042722.asp
From: CCP4
Rice
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu<mailto:pr...@uchicago.edu>
http://bmb.bsd.uchicago.edu/Faculty_and_Research/
http://www.rsc.org/shop/books/2008/9780854042722.asp
It might be that the protein is innocent and the FPLC is guilty. If FPLC pump
needs maintenance and is stuttering a bit when running at high pressure, the
salt gradient won't be as smooth as you'd like. If you have an ionic strength
detector, that trace will tell you. Otherwise, see if other
If you're only changing the 2-fold axis along c to a 2-fold screw axis, you
don't need to go back to the raw image files and reprocess them! Just tweak
the header of the .sca file and carry on (and take notes on what you did).
From: CCP4 bulletin board [CCP4BB@
Another annoying syndrome is people who download your coordinates, don't read
the paper, and then assume any insights are their own brand new ideas.
A biophysicist once said to me at a meeting "I hope you aren't offended that
we've found the DNA bend in your structure is flexible". The TITLE of
Something I recently learned from a friend: try changing the 4th base in the
coding sequence to G (e.g. ATGGxx...) and the ribosome will like it better. We
tried it recently on a frustrating construct and it worked like a charm.
From: CCP4 bulletin board [CCP4BB
MANY APOLOGIES the wrong address got auto-filled when I tried to forward this
to my lab.
Please delete.
++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu<mailto:pr...@uchic
; Robert Haselkorn; Geoffrey L. Greene; Sean Crosson;
Nancy Schwartz; Joe Piccirilli; Tao Pan; Keith Moffat; David R. Kovar; Tobin R.
Sosnick; Stephen C. Meredith M.D., Ph.D.; Phoebe A. Rice; Francisco Bezanilla;
Marvin W. Makinen; Stephen Kent; Donald F. Steiner Md.; Benoit Roux; D Allan
Drummond
Hi Gloria,
I've gotten useful results from these two threading / prediction servers:
Raptorx and Phyre (for low homology, I found the original version works better
than phyre2).
Phoebe
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecula
d on a gel, and it goes easy on the budget
as well.
Phoebe
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu<mailto:pr...@uchicago.edu>
http://bmb.bsd.uchicago.edu/Faculty
Check the pseudo-precession pics (0kl plane, etc). A rhombohedral crystal that
is indexed as P6xx may have seemingly-bizarre systematic absences that could
trick one into thinking its P63xx.
From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of
EEK, it seems to me that anything called simply "FP" should be unadulterated
FP. If the software modifies a column in some way, it should give it a new
label, shouldn't it?
++++++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biolog
red-green color blind?
thanks,
"Professor Rice"
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu<mailto:pr...@uchicago.edu>
http://bmb.bsd.uchicago.edu/Faculty_and
that in pymol.
So pymol developers, please consider yourselves wheedled ...
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu<mailto:pr...@uchicago.edu>
http://bmb.b
PMID:
10555960
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu<mailto:pr...@uchicago.edu>
http://bmb.bsd.uchicago.edu/Faculty_and_Research/
http://www.rsc.org/shop/bo
Looks like a typo to me: if you change the CRYST space group record from
P212121 to P21212, as the paper says it is, the packing problem goes away.
++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723
Among many possible reasons, the streaking could be caused mechanical stress to
the thin plates during mounting. Have you tried those loops that look like
miniature tennis rackets?
Phoebe
From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Rakesh
e point about Rfree holds for twinning or rotational
pseudo-symmetry: the Rfree flags should be expanded by the xtal symmetry
operators, not re-picked in the lower symmetry space group.
Phoebe
++++++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecu
try space group.
Phoebe
++++++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu
http://bmb.bsd.uchicago.edu/Faculty_and_Research/
http://www.rsc.org/shop/books/200
What happens if you solvent-flatten/flip/massage that map, but tell the
software the solvent content much lower than what you think it is now? Maybe
you'll find another copy of the molecule?
From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Andrey
end of the
glycosidic bond (note the atom number is different for pyrimidines vs.
purines). The sublime thing about W:C pairing is that it keeps the orientation
glycosidic bonds constant.
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The Univers
version (purified in a different lab)
doesn't.
thanks, Phoebe
++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu<mailto:pr...@uchicago.edu>
http://bmb.b
do with my time.
++++++++++
Phoebe A. Rice
Dept. of Biochemistry & Molecular Biology
The University of Chicago
773 834 1723; pr...@uchicago.edu<mailto:pr...@uchicago.edu>
http://bmb.bsd.uchicago.edu/Faculty_and_Research/
http://www.rsc.org/shop/books/2008/9780854042722.asp
__
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