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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: [log in to unmask]
To: [log in to unmask]
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.
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>
> 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:[log in to unmask]] Im Auftrag von LISA
> Gesendet: Montag, 17. Februar 2014 10:58
> An: [log in to unmask]
> 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
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