Dear Serah, It is likely that glycerol stabilized your protein via binding as it was available in plenty during purification. Therefore, you could attempt to purify the mutant enzymes in the presence of substrate or ligands. I am not sure if this will work, but worth a try since you have mentioned that "interferes with the binding of the substrates to my protein."
Good Luck, -Partha On Tue, Feb 23, 2010 at 3:07 AM, SERAH KIMANI <serah.kim...@uct.ac.za>wrote: > Dear all, > > Does anyone have an idea of something else that I can use instead glycerol > to maintain solubility of my protein? I have been having 10% glycerol in my > protein solution and this has helped me get crystals in like three different > conditions. However, I would want to get crystals of mutants-substrate > complexes, but unfortunately glycerol interferes with the binding of the > substrates to my protein. So, for the complexes to form, glycerol has to be > out (I have tested this using the wildtype enzyme both in the presence and > absence of glycerol). Now, in the absence of glycerol, I get a lot of > precipitation, and no crystals even after optimizing around the known > conditions with different protein concentrations. I have tried re-screening > for new conditions in the absence of glycerol, but I haven't found any > condition that yields crystals in the absence of glycerol. > > I was wondering if anyone might know of something else that I could use in > place of glycerol in my protein solution?? > > Regards, > > Serah > University of Cape Town > > ______________________________________________________________________________________________ > > > UNIVERSITY OF CAPE TOWN > > This e-mail is subject to the UCT ICT policies and e-mail disclaimer > published on our website at > http://www.uct.ac.za/about/policies/emaildisclaimer/ or obtainable from > +27 21 650 4500. This e-mail is intended only for the person(s) to whom it > is addressed. If the e-mail has reached you in error, please notify the > author. If you are not the intended recipient of the e-mail you may not use, > disclose, copy, redirect or print the content. If this e-mail is not related > to the business of UCT it is sent by the sender in the sender's individual > capacity. > > _____________________________________________________________________________________________________ > >
