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Dear Lisa, Good think is that your protein is crystallizing and diffracting X-rays. I am not sure if your protein is polymerizing. This could be a simple precipitation. Please check your pH, salt conditions or if you require some co-factors etc in addition to checking with bME as suggested by Herman. Anthony ----------------------------------------------------- Dr. Anthony Addlagatta Center for Chemical Biology Indian Institute of Chemical Technology [IICT] Tarnaka, Hyderabad AP-500 607, INDIA Tel:91-40-27191812 Web: https://sites.google.com/site/chembioliict/home/dr-anthony-addlagatta-1 ---------- Original Message ----------- From: [email protected] To: [email protected] Sent: Mon, 17 Feb 2014 10:06:18 +0000 Subject: [ccp4bb] AW: [ccp4bb] protein polymerization > *********************** > This message has been scanned by the InterScan for CSC SSM at IICT and found > to be free of known security risks. > *********************** > > Dear Lisa, > the first thing to check is whether the polymerization is due to disulfide > bond formation. If you run a gel of your protein with one sample boiled in the presence of b-mercapto ethanol and one sample boiled in the absence of bME, this should tell you whether disulfide links are the culprit. If disulfide links are the problem you could try adding DTT or TCEP to all your buffers, or to mutate the suspicious cysteines away. > Good luck! > Herman > > Von: CCP4 bulletin board [mailto:[email protected]] Im Auftrag von LISA > Gesendet: Montag, 17. Februar 2014 10:58 > An: [email protected] > Betreff: [ccp4bb] protein polymerization > > Dear All, > > My proein is polymerzated and elutate in the void volum when runnning gel > filtration Superdex 200. I can get a small crystal after a lot of optimization. But the resolution is still very low (about 8A ). I try to find the sites involved in polymerization. Is there some software to predicat the amino acids which are involved in protein polymerization? And All suggestion to the crystal optimization are welcome. Thank you. > > Best Regards, > > Lisa ------- End of Original Message ------- ------------------------------------------------------------------------------------------------- Note: The information contained in the e-Mail message and/or attachments to it may contain confidential or privileged information. If you are not the intended recipient, any dissemination, use, review, distribute, prinitng or copying of the information contianed in this e-Mail message and/or attachments to it are strictly prohibited. If you have received this communication in error. Please notify us by reply e-Mail or telephone and immediately and permanently delete the message and any attachment. Thank you.
