I use ccp4 to create Fo-Fc or 2Fo-Fc omit maps, then read them into Pymol
and display them with the isomesh command with an appropriate carve radius.
That might pick up an unmodelled blob.
__
Roger Rowlett
On Thu, Jul 18, 2024, 10:33 AM Andrea Smith
wrote:
> Hi,
>
>
&
ively
straightforward to infer hydrogen bonding interactions and poorly resolved
blobs that are likely loosely bound sulfate or phosphate.
Cheers,
Roger Rowlett
Gordon & Dorothy Kline Professor, Emeritus
Colgate University
Department of Chemistry
On Mon, Dec 25, 2023, 2:51 PM Tom Peat <
months.
Roger Rowlett
Gordon & Dorothy Kline Professor, Emeritus
Dept of Chemistry
Colgate University
On Mon, Oct 16, 2023, 2:52 AM Guillaume Gaullier <
guillaume.gaull...@kemi.uu.se> wrote:
> Maybe a naive question, and definitely going off-topic… but what if the
> successful candid
structure.
Roger Rowlett
Gordon & Dorothy Kljne Professor, Emeritus
Department of Chemistry
Colgate University
On Wed, Aug 10, 2022, 11:00 AM Thomas, Leonard M. wrote:
> Hello All,
>
> I have run into something odd. In working on a structure for one of the
> groups I work with regularl
Another possibility to model is a couple of S-hydroxy cysteines. We've seen
that in a Cys-amidase where there was artifactual oxidation damage to the
protein during storage or RT crystallization.
Roger Rowlett
On Wed, Dec 22, 2021, 8:00 AM Andrew Purkiss
wrote:
> As you say, the Cys
they were much more difficult for my
students to break, and they last for years. (My students never succeeded in
breaking one.) Gel-filled electrodes are garbage. Refillable double
junction electrodes are the way to go for stable readings, especially in
Tris, and minimizing heavy metal contamination.
of rate vs. substrate
data. For true product inhibition, there are several possible models that
could be considered, based on the behavior of the data. Consultation with
an enzyme kineticist might be warranted for for complex behavior.
Roger Rowlett
On Fri, Jun 18, 2021, 3:53 AM Harmer, Nicholas
wrot
Human carbonic anhdydrase II is very expressible in *E. coli,* and
purifiable in one step via affinity chromatography with
para-aminobenzenesulfonamide affinity resin (which is relatively easy to
make, and reusable for many years.) It can be assayed by stopped-flow
spectrophotometry for CO2 hydrati
Salt concentrations less than 100 mM can lead to nonspecific adsorption to
the gel exclusion media, potentially leading to band broadening, and
delayed elution. Overloading gel exclusion columns (more than 2-4% Vt) can
also lead to elution band artifacts. Check these issues first.
Roger Rowlett
Almost certainly a metal ion, possibly Ni(2+) if a Ni-NTA column was used
for purification. Ni-N bond lengths are typically around 2.0 A. Additional
density is probably coordinated water molecules, which should have similar
Ni-O bond distances around 1.9 A. It is fairly common to find adventitious
, https://sites.google.com/colgate.edu/xrd-protocols.
Roger Rowlett
On Tue, Jun 30, 2020, 7:25 AM Daniele Veggi wrote:
> Hi Yadav,
> yes, this is only a representation of what I would like to do. If during
> the refinement will continue to appear density, probably I'm on the good
&g
stive of a
thioketal adduct or a simple disulfide bridge. Building the thioketal, if
actually present, will require addition skill with ligand building and
Coot. You are essentially attaching an isopropyl group to the two Cys
sulfur atoms.
Roger Rowlett
Gordon & Dorothy Kline Professor, Eme
reasonable and space group was likely correct, and that I had the correct
number of chains per ASU. My initial MR attempts, based on Matthew's number
estimates, were 2 chains short, which was obvious from crystal packing.
Cheers, and good luck!
Roger Rowlett
Dorothy & Gordon Kline Professor,
S-hydroxycysteine can cartainly show up under nonreducing storage or
crystallization conditions, as an artifact.
Cheers,
Roger Rowlett
On Fri, Mar 27, 2020, 5:45 PM Chris Fage wrote:
> Hi Richard,
>
> I recently observed the sulfenic acid derivative of a cysteine residue
> (S
-click. Again, you have to go into the mouse configuration
settings.
I have two mice (a 3-button and a 2-button) configured to work properly
with Wincoot in Win10.
Roger Rowlett
Gordon & Dorothy Kline Professor, Emeritus
Colgate University
On Mon, Mar 23, 2020, 11:56 AM Schreuder, Herman
Chloride coordination spheres are typically tetrahedral (CN=4) or square
pyramidal (CN=5), and occasionally octahedral (CN=6). Arg and His and well
as amide nitrogens are common protein ligands, and it is possible but not
common to see carboxylates in the coordination sphere.
Roger Rowlett
On
?
Roger Rowlett
Gordon & Dorothy Kline Professor, Emeritus
Colgate University
On Wed, Mar 4, 2020, 12:20 PM Nukri Sanishvili wrote:
> Hi Jessica,
> You do not say how well is the rest of the structure refined.
> First, you should refine the structure best you can, without placing
>
. by manipulating protein/screen volume ratio.
__
Roger Rowlett
On Wed, Jan 8, 2020, 11:16 AM Armando Albert wrote:
> Dear all,
> I was wondering how to guess the optimal protein concentration for the
> initial crystallisation trials. Is there any trick or assay other
to simplify the search.
Good luck.
__
Roger Rowlett
On Wed, Nov 13, 2019, 10:33 AM Robert S Phillips wrote:
> I have been working on a protein structure which has been hard to solve by
> molecular replacement.
>
> Unit cell: (60.6, 172.34, 196.42, 90, 90, 90)
> Sp
The current version of CrysalisPro will directly import several image
formats, or you can convert images to Esperanto format to import. The
latter is a bit clumsy but does work.
__
Roger Rowlett
On Mon, Nov 4, 2019, 9:54 AM Almudena Ponce Salvatierra <
maps.fa...@gmail.com>
Won't this depend on the relative final concentrations of A and B in the
two experiments? If A going into excess B will have different mass action
considerations that B going into excess A. Even if the final concentrations
of A and B are stoichiometric, the initial stages of the titration will
have
numbers, though, so 2 is not a great improvement, e.g.
__
Roger Rowlett
On Thu, Aug 29, 2019, 1:42 PM Jonathan Cooper <
0c2488af9525-dmarc-requ...@jiscmail.ac.uk> wrote:
> It would be useful to know the number of molecules per asymmetric unit and
> the sequence simil
e are ways of using noncrystallographic symmetry
in your ASU to significantly improve your initial maps for rebuilding,
although sequence assignment may still be challenging at this resolution.
Roger Rowlett
On Mon, Aug 19, 2019, 8:15 PM Zhu Qiao wrote:
> Sorry for the initial message. I tried t
FYI,
When FFT is run in the CCP4i interface, the output file destination is
configured incorrectly. Examining the batch file that is created for the
job, it appears that CCP4i ignores the MAPOUT filename and substitutes the
file name of the temporary file, which is (alas) deleted in the last step.
Please find attached a description of a position opening at Colgate
University that may be of interest to a protein crystallographer interested
in working with undergraduates in a research-rich, research supportive
environment. Colgate is equipped with an Oxford Diffraction home source,
Gryphon dro
Question:
While it is perhaps understandable that reviewers may not always have
sufficient information to evaluate the quality of the interpreted model
(and must trust that authors have acted in good faith in representing
omit maps, etc.), is it not reasonable to expect reviewers to comment on
If it is as it appears, it is disappointing to see this in JACS. I would
expect better. Unfortunately, reviewers don't always get a lot of
information to judge quality of structures (which has been discussed
extensively on prior occasions on this board), so some trust is
involved that what dat
How about BioGel-A50m with an exclusion limit of 50 million Da? I've used
A15m for complexes as large as 500 kDa and it seemed to work well.
__
Roger Rowlett
On Wed, Apr 17, 2019, 11:11 AM Mohinder Pal
wrote:
> Dear all,
>
> I would like to gel filter a multi p
donor to acceptor
hydrogen bonding contacts. Sulfate rarely has donor to acceptor hydrogen
bonding contacts, as it is SO4= at any reasonable pH.
Roger Rowlett
On Sat, Feb 16, 2019, 4:06 AM 张士军 <21620150150...@stu.xmu.edu.cn wrote:
> Dear all
>
> I have got a crystal grown at the co
You might be able to distinguish sulfate from phosphate by examining
hydrogen bonding partners. Phosphate can donate one or two hydrogen bonds
at neutral pH values, whereas sulfate is usually only a hydrogen bond
acceptor. (Having said that, we have published a structure where a sulfate
clearly int
Iodine is ideally suited for Cu K-alpha SAD phasing, and iodide ions can
normally be easily added by soaking crystals in potassium iodide
containing solutions, which can be done at the time of cryopreservation.
A quick lit search will turn up the appropriate protocols. For
structural genomics w
-. Blob 2 is less likely to be metal around those Arg residues, but Cl-
or bicarbonate are possible and well-known in this kind of coordination
environment. Cl- is in your mixture, and bicarbonate can accumulate,
especially at alkaline pH from atmospheric CO2.
Cheers,
Roger Rowlett
On Oct 2
Besides empirically adjusting the weighting factor for X-ray data to
increase the geometric constraints, have you tried jelly-body refinement or
refinement with external constraints? The latter two methods can be helpful
when refining low resolution data.
Roger Rowlett
On Jul 12, 2017 7:33 PM
.
Cheers,
Roger Rowlett
On Jul 12, 2017 12:01 PM, "Alun R Coker" wrote:
> So, if we have a commercial 96 well screen where more than around 60% of
> the drops precipitate. It may be worth diluting the whole screen say (30ul
> screen and 20ul water in each well) and repeatin
We've had a Malvern Zetasizer for many years.
___
Roger S. Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346
tel: (315)-228-7245
ofc: (315)-228-7395
fax: (315)-228-7935
email: rrowl...@colgate.e
you will have lost a lot of data in the shadow
area, and the pin shadow is variable depending on the omega angle. If you
have crystals available, it would be worth collecting again properly.
Roger Rowlett
On Apr 27, 2017 5:18 PM, "Dr A.A. Jalan" wrote:
> Dear all,
>
> Thank y
;mailto:kell...@janelia.hhmi.org>> wrote:
A bit off topic, but I’ve always wondered how TRIS got so popular
what with it’s pKa of 8.3—does anyone know?
JPK
*From:*CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK
<mailto:CCP4BB@JISCMAIL.AC.UK>] *On Behalf Of *Roger Rowlett
*Se
What are you dialyzing against? Your storage solution should typically
be buffered away from the pI and contain at least a small amount of
kosmotropic salt, e.g. NaCl. Some proteins will require additional
stabilizing/solubilizing agents such as glycerol or reducing agents.
FYI, Tris-Cl, pH 7.5
Once again, chemical intuition may help. At neutral pH values, sulfate
is going to be present at SO4(2-), whereas phosphate will be present as
H2PO4(-) or HPO4(2-). The hydrogen bond network supporting binding may
be able to offer clues. Sulfate is not likely to have any H-bond
acceptors in its
. Bicarbonate ions will populate this site in crystals after
a few minutes soak at 100 mM concentration. The same crystals soaked in as
much as 1M (!) nitrate overnight do not bind nitrate ion in this site.
Roger Rowlett
On Nov 10, 2016 3:42 PM, "Keller, Jacob" wrote:
> Dear Crys
I've seen this kind of thing before.
Case 1: two crystal forms in the same droplet, C2 and C222. If you
looked closely, you could tell them apart and I was pretty good at
getting a high percentage of the desired space group by looking at the
crystal forms. The C2 form diffracted better, so I f
I'll also recommend Buccaneer. You might try using a combination of
PARROT for density modification and NCS averaging followed by
autobuilding with BUCCANEER using initial phases from your MR solution.
You only have two copies of the protein in the ASU, so you only get a
modest boost in electro
Except for graphics-intensive programs (e.g., Coot, Pymol), it is
possible to run Linux within a VM in windows, and you can even share
files with the other OS. I actually do it the other way 'round, running
Windows in Linux via wine to have a one-box solution for processing data
from our Oxford
If the metal center is a stable complex then ICP-OES or XRF (e.g. TXRF)
methods can easily identify and quantify metals present in a small sample
of protein.
Roger Rowlett
Dear All
I have solved a structure of a metal-ion dependent exonuclease enzyme. In
homologous structures, two or three
I think I would be tempted to chainsaw one of the ensemble chains of
2IT8 (they look very similar except for side-chain disposition) and use
1 or 2 of these as search models in a Phaser run. If this works, you
should see good Z-values and the final result inspected in Coot should
show good mole
g/content/276/5/3247.full
Shane
On Mon, Jun 22, 2015 at 2:15 PM, Roger Rowlett <mailto:rrowl...@colgate.edu>> wrote:
I agree...one possibility is a M-His(2)-Lys-(OH2) site. Possible metal
ions would include Zn(II), although Lys is a relatively rare ligand in
zinc-
g/content/276/5/3247.full
Shane
On Mon, Jun 22, 2015 at 2:15 PM, Roger Rowlett <mailto:rrowl...@colgate.edu>> wrote:
I agree...one possibility is a M-His(2)-Lys-(OH2) site. Possible metal
ions would include Zn(II), although Lys is a relatively rare ligand in
zinc-
I agree...one possibility is a M-His(2)-Lys-(OH2) site. Possible metal
ions would include Zn(II), although Lys is a relatively rare ligand in
zinc-metalloenzyme sites.
Cheers,
___
Roger S. Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colg
Appu,
You will have to edit your .tcshrc (startup) file to point to the
correct setup script with a line like this:
source /usr/local/xtal/ccp4-6.5/ccp4.setup-csh
If you are using a different shell, e.g. bash, you will have to edit the
appropriate startup file, e.g. .bashrc.
You will likel
If you mean generation of pdb coordinates of specific symmetry chains (not
just viewing) then you can do this with the symexp command in pymol. Select
the desired symmetry partners and save as pdb. You may want to edit
duplicate chain id labels in coot or a text editor.
Roger Rowlett
On May 22
Are the Cys residues in question on the surface of the protein? DMSO
(which is in the crystallization mix) is a weak oxidant and could
conceivably contain impurities like dimethylsulfide and methanethiol
which could form difulfides with surface Cys residues. Or there could
have been sulfides ca
Was the research publicly funded? If you receive funds from NSF, for
example, you are expected to share and "make widely available and
usable" software and inventions created under a grant (section VI.D.4.
of the Award and administration guide). I don't know how enforceable
that clause is, howe
In these coupled enzyme assays, concentrations of [E1], [S], [E2] and
[NADH] need to be chosen such that the rate of the second reaction is at
least one to two orders of magnitude faster than the first, otherwise
the measured rate -d[NADH]/dt will not be rate-limited by -d[S]/dt.
Normally this
I wouldn't mind knowing how to source this lamp as well. But FYI, the
lamps are usable for years after the FPLC gives you the dire "low
intensity" warning. We just ignore it until the lamp goes completely
dark or it's impossible to measure normal absorbances. We've used the
current lamp for man
We rarely use glycerol anymore, because it seems to fail so often for
many of our current proteins. Try glucose, 25-30%. This is most
conveniently done by weighing 125-150 mg of glucose in a microcentrifuge
tube, then addding well solution to the 0.5 mL mark and mixing until
completely dissolve
If your protease depends on an oxyanion hole for stabilizing the
transition state, fluoride ions are known to be a potent inhibitor of
these proteases. (It is a quasi-diagnostic test for serine-type
proteases, and related cysteine proteases.) This might allow you to get
reactant bound without c
On Apr 20, 2015 8:28 PM, "Roger Rowlett" wrote:
> Depends on what you want to accomplish... If you have a liter of crude
> lysate, "capacity" should be one of the choices. A step gradient is fast
> but low resolution; a gradient elution has more resolution but wi
rate and possibly capacity. Higher pump pressures are required to get
flow through columns packed with tiny particles sizes. To borrow an
analytical chemistry maxim: speed, resolution, capacity: pick any two.
Roger Rowlett
On Apr 20, 2015 6:45 PM, "xaravich ivan" wrote:
> Hi CCP4ean
The problem with crystallization is that is selects for the least
soluble, most packable species. Sometimes that works against what you
would like to know. That could include oligomerization state as well as
conformational state. For example, some of the allosteric carbonic
anhydrases stubbornl
In the CCP4i GUI you will find "Phaser Cell Content Analysis" under the
"Analysis" tab. IIRC, this calls up the Matthews Probability Calculator.
It will give you a good idea of the likely number of search models in
the ASU. For large numbers, e.g. >3, the most probable number is likely
to be un
sense to search for the domains separately? Did you truncate the model side
chains?
There are lots of ways to skin the cat depending on the problems
encountered. 3.0 A may be challenging for a low identity search model.
__
Roger Rowlett
Gordon & Dorothy Kline Professor
Depart
Bringing the pH closer to the measured pI would definitely help. The pI can
be measured on an IEF gel. Glycosylation, if applicable, could dramatically
increase solubility.
Roger Rowlett
On Feb 16, 2015 11:33 PM, "Mattiroli,Francesca" <
francesca.mattir...@colostate.edu> wrote:
Great reminder. But there are real non proline cis peptide bonds, including
highly conserved motifs in active sites, e.g. 3UAO and its homologs. I
would hope these don't get "corrected".
Roger Rowlett
On Feb 16, 2015 5:09 AM, "Tristan Croll" wrote:
> Dear all,
>
This is a known bug in Coot 0.7.x. You can add the metal ion correctly
using the Get Monomer dialog and selecting "CO" as the monomer. This is
fixed in Coot 0.8.1.
___
Roger S. Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colgate Universit
>> went for this space group well I would also try for the
other screw axes.
>> So should I Integrate the data from beginning with all
possible screw axes
>> of orthogonal space group? I am attaching the IDXREF.LP
screen shot he
Did you search all 8 possibilities of screw axes, e.g. P2221, P21212,
P212121, etc?
Roger Rowlett
On Jan 25, 2015 4:50 AM, "jeorgemarley thomas"
wrote:
> Hi, I have processed the data using XDS and space group found to be P 2 2
> 2 (16) and I used the phaser MR for first pha
The Bead Beater has a 15, 40, and 350 mL chambers. I haven't used mine
to homogenize yeast, but I suspect it is similar in performance to E.
coli disruption. (Different bead sizes are used for yeast than
bacteria.) We get excellent, gentle disruption of E. coli in 8 minutes
total. A French Pres
How you approach this will depend substantially on the sequence identity
between your target and your potential MR models. Definitely remove all
non-protein atoms from your search model, as these are not likely at all
to be present, or present at these positions, in your target. In
addition to
We usually try 25-30% glucose first. Weigh out 150 mg of glucose in a
microcentrifuge tube add well solution, dissolve, mix, and make up to
the 0.5 mL mark with well solution. If crystals crack in this solution,
make up a 40% glucose solution in mother liquor, and add this solution
in 0.25 drop
I use 3" clear duck tape. Amazon has it in packages of 6 rolls. Seems to
work OK, with maybe one well in a 2x96 well screen interacting with the
adhesive. I think it's an isopropanol or dioxane condition.
Roger Rowlett
On Dec 1, 2014 5:32 AM, "Mark J van Raaij" wrote:
I'm running home-built Ubuntu boxes with old, plain-vanilla CPUs (e.g.,
Q9300 or core i3/i5/i7) and 6-8Gbyte of RAM, and a cheap Nvidia video
card (e.g. GT 9xxx or GT 620).This is more than sufficient to do routine
structure solution. Any contemporary desktop or laptop computer should
be suffic
Adding metal ions will work properly through the "Get Monomer" dialog in
0.7.2. Until Coot is updated (either manually or through a new CCP4
release) this is a reasonable workaround.
___
Roger S. Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
There is a bug in version 0.7.2 Coot that causes metal ions added via
the "place atom at pointer" to be a water. However, if you add the metal
ions through the "Get Monomer" dialog I think it will work OK.
___
Roger S. Rowlett
Gordon & Dorothy Kline Professor
I've had a Gryphon for 2+ years and use it in an undergraduate
environment. It's been trouble-free, and there are no instrument
consumables, just blocks and trays. OK, I do have to feed it deionized
water and a PCR tube of diluted protein for each set. It can set a
96-well tray in under 2 minut
You've tried the obvious things. Lowering the IPTG concentration doesn't
really work all that well, at least in my hands. We find we can still
get maximal expression from many promoters like trc with only 0.2 mM
IPTG, and the response doesn't modulate well: it's either on or off.
Growing at low
The Art Robbins Scorpion has this function baked in the the operating
software. When I was shopping for liquid handling robots for
screen-making, only the Emerald Biosystems Opti-Matrix and the Scorpion
were affordable for a small lab. The Scorpion can handle more solutions
at one time.
_
I have experience with some proteins that don't tolerate freeze-thawing
very well. It's hard to say exactly what the physical chemistry of this
is, but it probably relates to (1) aggregation due to high concentration
or protein or salts during the freezing process as water is removed,
and/or (2
As always, look at the unit cell packing of your alternative solutions.
In all likelihood one of these two solutions from Phaser should pack
sensibly in the unit cell, and the other will not. You may get some sort
of quasi-reasonable-looking electron density out of the wrong solution
initially,
Define cheap. Several vendors offer SDS PAGE minigels for about $10 US a
pop. I get mine from Novex.
Roger Rowlett
On Aug 29, 2014 2:22 AM, "Theresa Hsu" wrote:
> Dear all
>
> Would anyone knows of source of cheap precast SDS-page gels?
>
> Thank you.
>
Excellent references. PEG 3350 appears to be hydrodynamically equivalent
to a 20 kD globular protein. So for efficient separation, your protein
needs to be significantly larger than 20 kDa on a GEC column. In a
centrifugal filter (which is very inefficient--you need many exchanges
and dilutions
Did you enter a valid pdb filename for a coordinate file when selecting the
option 'cover all atoms in pdb'?
______
Roger Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colgate University
On Aug 20, 2014 3:54 PM, "Wei Shi" wrote:
> Hi al
in the solution to minimize electrostatic
interactions, e.g. 100-500 mM NaCl.
Ultimately, your protein just may not be very soluble. That is potentially
OK...it will ppt and maybe xtallize well at low concentration.
Roger Rowlett
On Aug 19, 2014 1:52 PM, "Prashant Deshmukh"
wrote:
&g
degradation product of PEG200? Did any MPD get into the
mix? We have seen spurious oxidation of Cys to Cys-OH in thiol proteins,
e,g, PDB 3UAO.
Good luck. With that high quality ED, you'd think it would be easy to ID...
Roger Rowlett
Dear all,
We are currently working on a small GTPase
Harvey,
Depending on the zinc-binding site, it may not bind Fe(II) at all.
Zn(II) and Fe(II) have very different preferred ligand binding
environments. For many zinc-metalloenzymes, substitution with Fe(II)
would be difficult to impossible. In general, you will find it very
difficult to make
Exactly. Aimless will give you suggested resolution cutoffs based on CC 1/2
in the log file.
Roger Rowlett
On Aug 14, 2014 5:04 PM, "conan仙人指路" wrote:
> Hi Faisal,
>
> CC-half standard is valuable in evaluating the cut-off of highest
> resolution. Sometimes even if I/
takes a cup of
coffee, about 10-15 minutes to dispense. No way we're going back to hand
dispensing.
Roger Rowlett
Gordon & Dorothy Kline Professor
Colgate University
On Aug 5, 2014 5:05 PM, "Joseph Ho" wrote:
> Dear all:
>
>
> We are interested in purchasing ei
Increase the number of allowed clashes in Phaser, re-run it then look at
the packing of the solution found and identify the source of the
clashes. Possibilities for the clash issue include:
1. Wrong space group
2. Flexible loops or termini in search model not present or
differently arranged
po but could
substitute?
Thank you all so much for the help and advice.
-Adam
On May 19, 2014, at 12:55 PM, Roger Rowlett <mailto:rrowl...@colgate.edu>> wrote:
The answer depends on a number of questions:
* What metal ion are you trying to eliminate?
* What kind of metal-binding
The answer depends on a number of questions:
* What metal ion are you trying to eliminate?
* What kind of metal-binding site is involved?
o A peripheral or loose binding site? (e.g. surface calcium
ions)--these may respond to chelators
o An active site coordinated metal? (e.g.,
For P212121, I would put money on something divisible by 2 for the total
molecules per asu. Anything from 6-12 might be likely. One of the early
structures I worked on had 6 molecules per asu, which was darn near
impossible to find using momomers (at the time). The way it was
eventually solved
A logarithmic plot of cumulative entries to the PDB is approximately linear
and shows a growth rate of about 15% per year. That means it doubles in
size about every 5 years at current growth rate.
Roger Rowlett
On May 15, 2014 4:23 AM, "Colin Nave" wrote:
> From James's figure
See below.
Ceeers,
___
Roger S. Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346
tel: (315)-228-7245
ofc: (315)-228-7395
fax: (315)-228-7935
email: rrowl...@colgate.edu
On 4/17/2014 4:13 PM,
Try "yum install glib2-devel" (as root). I haven't used CentOS/Fedora in
a while, but I think this is the right package.
Cheers,
___
Roger S. Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346
A couple of thoughts:
* We have actually managed to grow quite a few crystals like this.
Sometimes they are not single crystals, but stacks of mis-aligned
plates that come apart easily when handled or subjected to osmotic
stress. Sometimes these stacks give great-looking diffraction
Assuming you can get the protein in solution, it should purify really
easily on a hydrophobic interaction column. When I was on sabbatical
leave at the NIH I helped a colleague purify a really hydrophobic,
aggregation-prone protein this way, and it was tantamount to affinity
purification. We us
How about a short swish in well solution + 25-30% glucose? Doesn't take
long to cryoprotect, just a quick sufrace coat. Sodium malonate? We just
froze some really fragile crystals from 1.8 M sodium formate in 3 M
sodium malonate and they held up really well. (Still didn't improve
their diffract
Your basic choices for protein assays are:
1. Alkaline copper methods (e.g., Biuret and micro-biuret)
2. alkaline copper + molybdate methods (e.g., Lowry, BCA assays)
3. Hydrophobic dye methods (e.g. Bradford)
4. UV methods (e.g., A280, A230, A210, etc.)
Method 1 is least sensitive to amino acid
Lots of choices. I usually try the crystallization solution + 30% glucose
first. Glycerol or ethylene glycol are other possibilities here. Another
possibility is 2.5-3.0 M sodium malonate at a similar pH.
Roger Rowlett
On Feb 6, 2014 11:40 PM, "Deepak Thankappan Nair"
wrote:
>
BeadBeater. http://biospec.com/. Gentle, aerosol-free way to break
15-350 mL of cell paste (2-150 g wet packed cells).
Cheers,
___
Roger S. Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346
te
, if you haven't already found
it, Hampton has a nice link to calculate volume of components
while designing a tray as long as you tell it the concentrations.
http://hamptonresearch.com/make_tray.aspx
Nick
From: Roger Rowlett mailto:rrowl...@colgate
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