And what are you dialyzing it against? JPK
From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Akilandeswari Gopalan Sent: Wednesday, March 29, 2017 9:38 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] protein precipitation reg Dear all, I am a PhD student doing structural studies on a few proteins from Mycobacterium tuberculosis. The gene encoding the proteins I work on are cloned into pet22b with c terminal His tag. the proteins are expressing well. upon purification I am getting good yield of protein but during dialysis, the proteins precipitate. Kindly suggest some solutions to avoid aggregation. pI of one protein is 9.7 and that of the other is 5.6 I am using 25mM Tris pH 7.5 and 100 mM NaCl buffer with 5mM beta-mercaptoethanol and 0.5% triton x 100 for lysis, the same buffer with 20-30mM imidazole for washing and 300mM imidazole for eluting the proteins. Thank you Regards Akila -- Akilandeswari G
