Yes, woops! I over-simplified. The distribution of <|I-|> is not
Gaussian, nor is it centered at zero. It is the distributions of I-
and that are Gaussian and centered at zero.
I also agree that the reason for the instability is because the "true"
value of I is zero. In fact, (at least in my
>For gel-shift assay, do people normally use a special gel tank for heavy
>metal work?
We used to use a Pharmacia Phast system, as this is bufferless and made it
very easy to contain heavy metal contamination. Ours caught fire and died a
couple of years ago, and I have yet to find a good cheap r
James,
Graeme is right. While does indeed (approximately) follow a
Gaussian, <|I-|> cannot. The absolute value operator keeps it
positive (reflects the negative across the origin), and hence it is a
half Gaussian. Its mean cannot be zero unless the variance is zero.
For standard norm
The Schwartz laboratory at MIT, Department of Biology (Cambridge MA,
USA) seeks to recruit an outstanding postdoctoral scientist in
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James,
I'm not sure you're completely right here - it's reasonably
straightforward to show that
Rmerge ~ 0.7979 / ()
(Weiss & Hilgenfeld, J. Appl. Cryst 1997) which can be verified from
e.g. the Scala log file, provided that the *unmerged* I/sigma is
considered:
http://www.ccp4.ac.uk/xia/rmerge
I tried plugging I/sigma = 0 into your formula below, but my calculator
returned ""
-James Holton
MAD Scientist
Graeme Winter wrote:
James,
I'm not sure you're completely right here - it's reasonably
straightforward to show that
Rmerge ~ 0.7979 / ()
(Weiss & Hilgenfeld, J. Appl. Cry
Hi Laurie,
An related, but off-topic comment follows.
This particular issue is well suited to the single crystal
spectrophotometer that we have integrated into beamline X26-C at the
NSLS. We have observed that many heme protein crystals change either
redox state and/or ligand state as a functi
Actually, if I/sd < 3, Rmerge, Rpim, Rrim, etc. are all infinity.
Doesn't matter what your redundancy is.
Don't believe me? Try it.
The extreme case is I/sd = 0, and as long as there is some background
(and, let's face it, there always is), the "observed" spot intensity
will be equally lik
Hi Frank,
Off from the original topic but important to clarify. If I misled the
concepts, I apologize.
Outer shell Rmerge will always be very high:
--
True! Especially when I/Sig ~ 1 or less.
Only I/sigI (and completeness, although it's related) is really
relevant for deciding
Hi folks,
Pfizer protein folks taught me a useful trick: the 1L centrifuge rotor
liners from Beckman-Coutler (catalog number 369256 if you're curious).
They are quite awesome in the sense that instead of scraping biomass out
of the centrifuge tube one can instead heat-seal and freeze the lower
por
Hi,
1. KSCN will suck up a lot of the Hg and other metals -- can you switch to
something else?
2. Hg typically does not enter the protein core.
3. It's generally not necessary to use a special tank, as long as you
treat the resulting solutions as heavy-atom waste; but of course each
lab's safety p
There is a Xenon chamber at 14-1 if I remember it correctly.
I never had luck though.
How about Iodine or Gadolinium?
Jürgen
..
Jürgen Bosch
Johns Hopkins Bloomberg School of Public Health
Department of Biochemistry & Molecular Biology
Johns Hopkins Malaria Research Institute
I think it is up to the buyer to gain knowledge about prices from other
companies. So I agree the survey on CCP4 was a good idea, but we should
stop the "legal" discussion here (or continue outside ccp4) - it's not
really crystallography :-)
Clemens
> -Ursprüngliche Nachricht-
> Von: CCP
Xenon/Krypton, anyone? If you have the equipment, might as well try it. I
think I have seen the apparatus at some beamlines, although I did hear
recently that xenon is ridiculously expensive now.
Jacob Keller
***
Jacob Pearson Keller
Northwestern Univers
I think that since there is not a uniform price, competition is not playing
a significant enough role in pricing. So I ask: why is competition not
playing a role? And I would suggest: perhaps the bigger companies are
exploiting the lack of knowledge about the true market price of IPTG.
Happily,
Totally agree with what you write.
Yet, I was only answering the question asked (that dealt mostly with
"van Der Waals contacts and effective contacts").
At a time of crisis (DeltaG = only a few kcal/mol), I would say 0.6 kcal/mol
would be a little more than negligible. Question of balance.
Nadir
X Xiong, Cellular & Molecular Medicine wrote:
My Question is:
Does mercury tends to get into the protein core to denature protein or not?
This is more likely to happen for a small "bare" Hg like Hg2+ in HgCl2
or Hg(OAc)2 than it is for a large organomercury compound like PCMB,
PCMBS etc so i
It might be worthwhile to consider the energy column in the pdf:
At RT we have about 0.6 kcal/mol thermal energy, so a
*single attractive* vdW interaction has little impact -
it is generally the sum of many of those contributing to
notable and important attractive forces.
For a *single repulsive*
Dear All,
We have a very similar problem, I have got a protein of 200 residues
predicted to have two free cys (but 14 internal cys!), 1 free histidine, 2
methionines (all predicted to be free).
Native crystals diffracted to 2.2 A was obtained in 200 mM KSCN, 20% PEG
3350, 13% glycerol, bis t
When refining a heme protein in Refmac5, if one uses HEM.cif, what is
the default oxidation state of the Fe? Does one get residual density
if it is specified as Fe3+ but it's really Fe2+? Or does one infer
from the ligand bound that it's one or the other? WE have a hme
protein that was o
I would also include a NaI and NaBr quick soak in 250 mM, SCN- is
considered a pseudohalide
cheers Preben
Sebastiano Pasqualato wrote:
Hi all,
I've got crystals of a protein of ca 200 residues, with 2 free
cysteines, 5 histidines, 2 methionines.
We have nice diffraction for the native crystals
Hi Sebastiano,
In addition to the heavy atom database server and the suggestions made
here, you could try the strategy described in this paper:
Lott JS et al. (2003) Acta Cryst D Biol Crystallogr, Dec (Pt 12):
2242-6. Making the most of two crystals: structural analysis of a
conserved hypot
Sebastiano--
http://www.sbg.bio.ic.ac.uk/had/
good table of HA and pH ranges
http://www.shapirolab.org/lab-links/had.html
links to a variety of HA databases
HTH!
annie
Annie Hassell
Glaxo Smithkline
5 Moore Drive
RTP, NC 27709
919/483-3228
919/483-0368 (FAX)
annie.m.hass.
Dear Sebastiano,
why not have a look at
http://sis.niaid.nih.gov/cgi-bin/heavyatom_reactivity.cgi.
A reprint of the original paper can be found at
http://sis.niaid.nih.gov/pub_pdf/Agniswamy-et-al-2008.pdf
We did not specifically look at Bis-Tris Propane but did look at Tris pH 8.5
and H
Dear All,
I never tried it myself but could it be a sulphur SAD/MAD a possible
alternative?
In my experience with heavy atoms (luckily just 2 proteins!) you may
waste all of your crystals without finding any heavy metal bound! Or may
be the first one you try works
Ciao
Stefano
-Origina
Hi,
Mercury will likely form a precipitate with you SCN- anions...
solubility in water is 0.6 g/L (about 3 mM) at 25°C...
Since you are not in water, at less than 25°C, and with you high
concentration of SCN- ions, don't expect to have high concentrations of
*soluble* Hg++ in you medium. Prolo
Whps!
Watch out for problems with your *HIGH* pH, though. Some salts will form
insoluble hydroxide salts at pH 8.8. (HgCl2 & K2HgI4 should be okay).
2009/7/15 Sebastiano Pasqualato :
> Hi all,
> I've got crystals of a protein of ca 200 residues, with 2 free cysteines, 5
> histidines, 2 methi
Hi Sebastiano,
Given that your crystallization condition contains KSCN, I would certainly
give Hg(SCN)2 a go among other Hg options for a protein containing free
cysteines. Hg(SCN)2 might end up being more compatible with your crystal
form, and might thus minimize non-isomorphism issues.
Best of
Hi Sebastiano,
Free Cys' are crying out for Mercury derivatives. Try 2-3 with varying
sizes of additional groups - HgCl2, K2HgI4, PCMB, from the "Magic
seven" would be a good place to start.
Watch out for problems with your low pH, though. Some salts will form
insoluble hydroxide salts at pH 8.8
Hi Sebastiano,
Have a look at HATODAS:
http://hatodas.harima.riken.go.jp/
Good luck,
Miguel
Le 15 juil. 09 à 14:33, Sebastiano Pasqualato a écrit :
Hi all,
I've got crystals of a protein of ca 200 residues, with 2 free
cysteines, 5 histidines, 2 methionines.
We have nice diffraction for
Hi all,
I've got crystals of a protein of ca 200 residues, with 2 free
cysteines, 5 histidines, 2 methionines.
We have nice diffraction for the native crystals, that grow in 150 mM
KSCN, 17% PEG 3350, bis tris propane pH 8.8.
We are crystallising the SeMet derivative, but I'm not completely su
Fermentas 25g 400$ (Dioxane-free)
Jürgen
On Jul 15, 2009, at 8:01 AM, Clemens Steegborn wrote:
Hi Jacob,
I am not sure I understand why a spread in pricing indicates illegal
activity.
Wouldn't uniformly high prices indicate something illegal, and very
different
prices just indicate you have a c
Hi Jacob,
I am not sure I understand why a spread in pricing indicates illegal
activity.
Wouldn't uniformly high prices indicate something illegal, and very
different
prices just indicate you have a choice of suppliers, qualities etc?!?
And just to mention two more supplier names (again: differen
Dear Wei,
The unit-cell dimensions are certainly not excessive, we recently published the
2.8A 27kDa uridylate kinase structure from B. anthracis whose crystals were
P6122, a=b=87, c=384 with three UK in the ASU.
As you are apparently not seeing any self rotation peaks, have you looked for a
t
I'm not clear what is the reasoning here behind using a low res cutoff for the
SRF (i.e. lower than the data limit). It seems to me that using all valid data
available can only increase the signal/noise ratio, and omitting good data can
only have a deleterious effect (as will any kind of data i
Um, can't resist here (although not directly relevant to the original
question):
1) Your dataset has a high overall Rmerge. The outmost shell (70%) is
very high, which suggests a need to shrink resolution. What about
I/s(I), redundancy and completeness? Also, how many
reflections (percentage
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