Re: [ccp4bb] Regarding quality of protein for crystallization

2010-09-26 Thread Artem Evdokimov
1. your information is helpful but not enough to tell if this double-band pattern is a product of e.g. proteolysis or of abortive translation. You don't specify what these 'minor differences' are - in the world of MS there is no such thing as 'minor' since every difference is either a significant o

Re: [ccp4bb] problem in heavy metal soaking

2010-09-26 Thread Paul Smith
Hi Seema, In theory, Ammonium Sulfate contains some small fraction of neutral ammonia which can act as a strong nucleophile and react with many heavy metal compounds. That being said, I recently phased two structures with mercury soaks, both of which contained ammonium sulfate. The first was a

[ccp4bb] Home Source Options

2010-09-26 Thread Paul Smith
I'm interested in opinions/advice on home source systems. As synchrotron beamlines are more powerful and more accessible than ever, a home source is really only needed for crystal screening, if at all. With that idea in mind, what are options out there for buying and running a home source in th

Re: [ccp4bb] Regarding quality of protein for crystallization

2010-09-26 Thread Daniel Bonsor
I assume that the loss of the peptides that you observe for the smaller protein is at the other terminus from the tag? If it is at the terminus where the tag was it would suggest that removal of the tag using proteolysis is the most likely cause. Though saying that Factor Xa/thrombin is not 100%

Re: [ccp4bb] Regarding quality of protein for crystallization

2010-09-26 Thread Jürgen Bosch
What are the minor differences in peptides ? Any PTM's perhaps ? Jürgen - Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry & Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Phone: +1-410-614-4742 L

[ccp4bb] Regarding quality of protein for crystallization

2010-09-26 Thread vikrant saa
I am working on 45 kd protein (pI 5.3) for crystallization . It is in pGEX vector.I do the expression (induction 24 degree 16 hrs) and purification from  Rosetta 2 DE3 cells (has additional rare codon).when I run the affinity chromatography purified protein (cleaved protein after removal of tag

[ccp4bb] please disgard

2010-09-26 Thread Bernhard Rupp (Hofkristallrat a.D.)
test of subscription change

Re: [ccp4bb] problem in heavy metal soaking

2010-09-26 Thread Patrick Loll
On 26 Sep 2010, at 9:00 AM, Seema Nath wrote: > I'm working with a protein which crystallizes in a mixture of PEG6K with 0.2M > AmSO4,my question is if there's any problem if I want to soak heavy metal > derivatives in this crystallizing condition? Does AmSO4 interfere in > heavy-metal soaking

[ccp4bb] problem in heavy metal soaking

2010-09-26 Thread Seema Nath
I'm working with a protein which crystallizes in a mixture of PEG6K with 0.2M AmSO4,my question is if there's any problem if I want to soak heavy metal derivatives in this crystallizing condition? Does AmSO4 interfere in heavy-metal soaking ? if yes, what's the reason? Thank you in advance.