Hi Xiaopeng
To add to Artem's comments:
Does the presumed gsh make a mixed disulfide in the active site?i.e. is it
covalently bonded to the active site via a s-s bond?
If yes then MS on your purified sample should easily give you the answer.
If a mixed s-s is indeed the scenario then purifying
Dear all,
I was working with a protein which is known to bind zinc. I tried to make a
limited proteolysis (with trypsin) after purification (metal affinity, ion
exchange and gel filtration; last step uses EDTA to remove bound metal ions)
in the presence and absence of zinc ions and I was quite
To add more information:
The proteolysis buffer was 50 mM Tris / HCl pH 8.0, 150 mM NaCl, 0.5 mM ZnCl
and 0.1 mM TCEP; protein concentration was ~ 25 µM. Proteolysis was carried
out at 4°C over 2 hours.
Thank you very much for the literature, Mark - I'll look into it.
Greg
2011/3/16
Hi Greg,
I am not sure why you are so surprised! If the zinc is altering the
conformation and/or folding of your protein, this might change the
accessibility of trypsin cleavage sites, thus changing your limited
proteolysis pattern.
Eg:
Metal-ion induced conformational changes in alkaline
I guess the only real choice is P2 21 21 or P21 21 21 - the absences
alng h00 could be a result of the pseudo-translation.
I cant explain the score - maybe there is something in the documentation?
But I am afraid after refinement in P212121 the resultant model is sure
to give the best score
MAR describe their latest Image Plate as a CCD. Their early Image Plate designs
were described as barium halide phosphor doped with Eu2+. Does anyone know why
they have kept the name Image Plate when everyone else calls it a CCD?
Rex Palmer
Birkbeck College
Maybe because a CCD detector still requires a phosphor layer to detect X-rays?
Tim
On Wed, Mar 16, 2011 at 12:59:14PM +, REX PALMER wrote:
MAR describe their latest Image Plate as a CCD. Their early Image Plate
designs were described as barium halide phosphor doped with Eu2+. Does anyone
check
Dear all,
Recently, I came across an obstacle on the purification and acitivty
measurement of my protein. My protein was expressed with an C terminal His
tag in the HEK 293T cells and purified by nickel affinity, anion
exchange and size exclucion chromatography. For every purification step, I
Hi everyone,
I recently purchased a 4L Vapor Shipper (Taylor-Wharton) from VWR. Turns out
it's designed for samples kept in canes, I wanted one that could hold pucks for
APS data collection. VWR won't take it back because I got rid of the
packaging. Please see description on the website for
I was very relieved to learn that my friend and colleague Hideaki Niwa who took
his MSc and PhD with me at Birkbeck is safe and well in Japan.
I believe that International the Red Cross is doing great work out there and
need all the help they can get. You can donate by going to the link
I guess it depends on what your activity is. Can you divulge that?
Could it be that Ni is necessary?
Jacob
On Wed, Mar 16, 2011 at 11:28 AM, Harvey Rodriguez
h.rodriguez.x...@gmail.com wrote:
Dear all,
Recently, I came across an obstacle on the purification and acitivty
measurement of my
Depending on what the expected activity is, its worth considering the
highly-depressing possibility that the activity seen in the impure sample was
due to impurities: for example, barely-visible-on-a-gel chaperones can give a
nice ATP hydrolysis signal, and DNA ligases float about with an AMP
Hi Harvey,
Well, knowing nothing about your protein, allow me to ruminate anyway...
It sounds like you are exploring the possibility of a metal ion or
other cofactor being lost. This is a reasonable first thing to check,
but your buffer exchange steps should allow small cofactors (smaller
Hi Harvey,
Could it be that the activity you're measuring comes from a contaminant?
Did you test the other fractions from SEC or IEX?
Cheers,
Alex
2011/3/16 Harvey Rodriguez h.rodriguez.x...@gmail.com
Dear all,
Recently, I came across an obstacle on the purification and acitivty
Le 16/03/2011 17:59, REX PALMER a écrit :
Would it be possible to get information through the CCP4BB about
colleagues who do not answer mails ?
I'd like to have news about TAKENAKA Akio, Faculty of Pharmacy, Iwaki
Meisei University, Tokyo Institute of Technology.
Thank you if somebody can
http://japan.person-finder.appspot.com/?lang=en
The latest numbers I read is that they have 140,000 records, unfortunately
there is no information about TAKENAKA Akio available yet.
Best wishes,
Thomas
On Wed, Mar 16, 2011 at 21:17, Philippe DUMAS
p.du...@ibmc-cnrs.unistra.frwrote:
Le
Thanks for the concern. I have replied to Prof. Dumas some minutes ago,
but should have done to CCP4BB (Thanks a lot for letting us to use
CCP4BB in this way. I think this to be an exceptional use of CCP4BB
under the catastrophic circumstances in Japan.). I heard from one of
his
Thank you to everyone who replied. I went through all the suggestions and
in the end used Jason's PyMOL script, using Thomas' cpv.distance suggestion
(which did make it much faster for me) and a few more modifications to
eliminate redundant pairing listings.
Bellow is the modified script, saved
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