Hi All,
A truly herculean response! Thanks everyone, I will process all of the
information and come up with a strategy.
Rhys
Hi Rhys,
It's worth paying close attention to your crystallisation conditions as
well - some heavy atom compounds will not be at all soluble in very
alkaline (they'll form insoluble hydroxides) or phosphate/sulphate
containing mother liquors.
A very low pH may reduce the binding efficiency of
Hi John,
Another way to screen for mercury derivatives.
Rachelle
vincent Chaptal vincent.chap...@ibcp.fr wrote:
Hi Rhys,
you already have a lot of suggestions to try. We all have our own reciepe for
good derivatization, and this is due to the fact that we don't really
understand what is
A favorite resource is Bart Hazes' web page on heavy atom derivatives.
http://homepage.usask.ca/~pag266/bart-hazes.html
Ho
Ho Leung Ng
University of Hawaii at Manoa
Assistant Professor, Department of Chemistry
h...@hawaii.edu
free cysteines? - pCMB
phosphate-binding? - tungstate
Best,
Matthias
-
Dr. Matthias Zebisch
Division of Structural Biology,
Wellcome Trust Centre for Human Genetics,
University of Oxford,
Roosevelt Drive,
Oxford OX3 7BN, UK
Phone (+44) 1865 287549;
Fax
You don't mention the condition, which is important (esp pH). Best pH values
are 6-8.
From personal experience: try K2PtCl4 and OsCl3.
Try ~5 mM for 30-60' (quick soak).
The advantage of osmium salts is that they give a nice color to your crystals
so you know something is binding.
HTH and GL,
There is quite a bit of literature on this, but my favorite paper is this:
http://www.ncbi.nlm.nih.gov/pubmed/18391402
Towards a rational approach for heavy-atom derivative screening in
protein crystallography.
Spoiler: The overall winner is ethyl mercury phosphate (Figure 7). Of
course,
More references to consider…
You asked about soaking times - here are two articles advocating quick soaking
at relatively high heavy atom concentration, which has worked well for us.
We've had good luck with thimerosal.
Acta Crystallogr D Biol Crystallogr. 2002 Jul;58(Pt 7):1099-103. Epub
And quick iodide soaks may be useful in the 200 mM range. See the sped-up
video:
http://www.youtube.com/watch?v=45Qc3jOPaKY
Quiz time: What wavelength would give iodide a similar signal to that of
selenium? Can one get a better signal than selenium by choosing a different
wavelength for
On Jan 15, 2014, at 10:27 AM, Jim Pflugrath wrote:
Quiz time: What wavelength would give iodide a similar signal to that of
selenium? Can one get a better signal than selenium by choosing a different
wavelength for data collection?
I'll bite,
At ~11,000eV Iodine has about 3.8
Hi
do not forget the clusters like Ta6Br12 or the lanthanides.
in case your interesting protein is a membrane protein there are some
choices that might work better than other
we have described it here.
http://www.ncbi.nlm.nih.gov/pubmed/16855303
This is not exclusive to membrane proteins at
Dear Rhys,
an important addendum to the magic triangle is the fact that it gives
you direct evidence whether or not the substructure search has succeeded
(by locating the triangle in the substructure solution) so that you can
carry on with phasing via density modification. At 3.5A resolution this
Keith's favourites over many years (shared with Gideon):
K2PtCl4
K2PtCN4
KAu(CN)2
Ethyl Mercury Thiosalicylate (gentle Hg reagent)
Try for about 2-12 hours at 1 mM initially. If crystals crack, reduce
concentration - at least they bind!
If these don't work, then finding a derivative is
Also, if you have a sample changer for easy screening, try 1 mM, 5 mM 20 mM of
each
for 1h, 5h, overnight, and screen all ~60 crystals for diffraction
overnight.Send those that survive
to the synchrotron.
K
On 15 Jan 2014, at 17:18, RHYS GRINTER wrote:
Hello message board,
My group has
Hi Rhys -
Don't forget to try sulfur-SAD, especially the multi-crystal version
published recently:
http://journals.iucr.org/d/issues/2013/07/00/ba5189/index.html
This seems well suited to your situation.
- Matt
On 1/15/14 12:18 PM, RHYS GRINTER wrote:
Hello message board,
My group has
Did anybody mention native gel electrophoresis to select suitable HA ions?
Worked for us really nicely in a situation were speed was essential.
Here's a reference: PMID 14646083
Klaus
===
Dr. Klaus Fütterer
Room 717,
Hello Rhys Grinter, hello to the ccp4bb community,
I don't necessarily want to advertise here one of the major research
topic of Eric Girard and late Richard Kahn lab (my previous lab ;-) ) but...
You could maybe try lanthanide complexes ?
They are composed by a chemical ligand, which can
Dear Rhys,
You may consider Xenon derivative, which could be prepared simply pressurizing
the protein crystals in a xenon chamber. It does not require any modification
of mother liquor. It just needs cryo-protectant where crystals are stable for
at least one to two mins. Higher Pressure (20
You may be missing a trick by not using metals as crystallisation additives if
your best
diffraction is only 3.5 Angstroms. Uranium has to be my favourite heavy metal,
even if
I've solved more structures with Pt and Hg. It gave crystals diffracting to 1.2
A from a
protein that otherwise gave
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