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 the
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 sur
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 Matthias
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 phosph
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 in
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
pres
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
wrote:
> Dear all,
>
> Recently, I came across an obstacle on the purification and acitivty
> measurement of my protein. My protein was
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 c
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
> Dear all,
>
> Recently, I came across an obstacle on the purification and acitivty
> measurement of my protein. My pr
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 tran
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
wrote:
> Le 16/03/2011 17:59, REX PALMER a écr
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 ex-students,
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 a
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