Re: [ccp4bb] suitable buffer for CD studies

2013-03-20 Thread Vitali Stanevich
Harsh, This article describes common buffers for CD on page 2: http://www.ncbi.nlm.nih.gov/pubmed/17406547 Or this article on page 8: http://www.ncbi.nlm.nih.gov/pubmed/16027053 It seems like phosphate is the best, because it has low absorption at 180-200nm region. From organic buffers

[ccp4bb] off-topic: detergents for the stabilisation of water-soluble proteins

2012-10-12 Thread Vitali Stanevich
Hi, Sorry for off-topic question. Does anyone have experience of the stabilisation of water-soluble proteins by detergents? Protein I'm working with is definitely water-soluble and has high yield, but, unfortunately, not very stable. Especially during concentration. So, we thought that adding

Re: [ccp4bb] Poor baculovirus stability at 4C?

2012-09-27 Thread Vitali Stanevich
Alexander, We produce baculovirus at Gibco SF900-II + 10% heated FBS + 1xPSG and then store at 4C routinely. And it seems to be quite stable. We have a virus that had no change for more than 2 years: we judge by the volume of it needed to infect_cells/produce_protein, no titer measurements. We

Re: [ccp4bb] off topic: reduced glutathione interfering with protein activity?

2012-08-29 Thread Vitali Stanevich
? Thanks, Peter On Wed, 29 Aug 2012, Vitali Stanevich wrote: Peter, Such high concentration of GSH may change the pH according to our experience. We usually use 50 mM Tris pH=8.0, 5 mM GSH, 3 mM DTT - so that you can load the sample on ion-exchange column after elution. If your protein

Re: [ccp4bb] Off-topic His-Antibody

2012-06-25 Thread Vitali Stanevich
The one we have is rubbish. I can tell a vendor name if you want off-list. A lot of non-specific bands + high background level makes it almost impossible to use against cell lysate. It sort of works against purified His-tag protein, but not perfect either. On Mon, Jun 25, 2012 at 4:33 PM, D

Re: [ccp4bb] Off topic about application of detergent on non-membrane protein crystallization

2012-06-15 Thread Vitali Stanevich
Donghui, There is detergent screen from Hampton: http://hamptonresearch.com/product_detail.aspx?cid=1sid=39pid=31 We set up detergent screen for crystal conditions we'd like to improve: 200nl(buffer) + 200nl(protein) + 40nl(detergent). Occasionally (very occasionally) it works, but don't put to

Re: [ccp4bb] SUMO(ULP-1) protease

2012-05-24 Thread Vitali Stanevich
Gloria, Why don't you introduce TEV-cleavage site? You can clone it between His- and sumo-protein_of_interest (if you want co-crystallise sumo with your protein) or between His-sumo and protein_of_interest (if you want to crystallise your protein only). Vitali On Thu, May 24, 2012 at 2:41 PM,

Re: [ccp4bb] Sequence Alignment Question

2011-08-18 Thread Vitali Stanevich
You can try this site: http://espript.ibcp.fr/ESPript/cgi-bin/ESPript.cgi If you have .pdb for one of your sequences, you'll be able to show secondary structure also. vitali On Thu, Aug 18, 2011 at 6:31 PM, Yuri yuri.pom...@ufl.edu wrote: Hello Everyone, A little off topic but, what is a