Jan, If the Cys residues are accessible, you could try DTNB to quantify the number of Cys, thus determining if they are reduced or bridged.
http://en.wikipedia.org/wiki/Ellman's_reagent Kelly ******************************************************* Kelly Daughtry, Ph.D. Post-Doctoral Fellow, Raetz Lab Biochemistry Department Duke University Alex H. Sands, Jr. Building 303 Research Drive RM 250 Durham, NC 27710 P: 919-684-5178 ******************************************************* On Wed, Jul 11, 2012 at 11:08 AM, Jan Rashid Umar <[email protected]>wrote: > Dear all, > > I am working on a protein where I have to stabilize the closed > conformation of the protein using disulphide bond. The strategy to design > the cysteine mutants is based on the molecular dynamic simulations, and > accordingly the residues were chosen. The ultimate goal is to trap the > ligand in closed conformation of protein and crystallize it. I am facing > few issues: Is there some reliable assay that can check the formation of > disulphide bonds in protein. Additionally, does anybody knows another > method(s) that can be used to trap a closed conformation. I look forward > for your suggestions and discussions on this issue. > > Thanks very much! > > Jan >
