Dear All, I have a 50kDa protein that is soluble and monodisperse at up to approx 1mg/ml (after Ni-affinity and size-exclusion chromatography).
However, it aggregates (probably both via disulphides and via 'sticky/hydrophobic patches') when I concentrate it towards 2-3mg/ml, even in the presence of several detergents. I don't want to add DTT since my protein should have several intramolecular disulphides....although I do have 2 free Cysteines, partially exposed. I have already tried mutating the Cysteines, with little improvement. Any suggestions for obtaining 5-10mg/ml? Does anybody have good experiences with usin L-Arg and L-Glu (e.g. At 50mM) to aid concentrating (as in the Golovanov AP paper, JACS, 2004, pages 8933...) Thanks for any input! Yours, Matt ==================================== Matthew J. Bottomley, Ph.D. Senior Research Biochemist IRBM / MRL-Rome ==================================== Notice: This e-mail message, together with any attachments, contains information of Merck & Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates (which may be known outside the United States as Merck Frosst, Merck Sharp & Dohme or MSD and in Japan, as Banyu - direct contact information for affiliates is available at http://www.merck.com/contact/contacts.html) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system.
