title: Three-state kinetic analysis of Chinese hamster dihydrofolate reductase unfolding by guanidine hydrochloride.
abstract: The unfolding behavior of dihydrofolate reductase from Chinese hamster in solutions of guanidine hydrochloride (GdnHCl) was studied. The GdnHCl-induced unfolding of the dihydrofolate reductase monitored by intrinsic fluorescence shows a biphasic transition, while the change in the enzyme activity is a single exponential process. The rate constant of inactivation is consistent with that of the fast conformational change. Therefore, the kinetic intermediate of protein unfolding should be a partially folded and inactive form. On the basis of the kinetic equation of substrate reaction in the presence of GdnHCl, all microscopic kinetic constants for the free enzyme and enzyme-substrate complexes have been determined. Both substrates, NADPH and 7,8-dihydrofolate, protect dihydrofolate reductase against inactivation.
authors.affiliation: National Laboratory of Biomacromolecules, Institute of Biophysics, Academia Sinica, Beijing, People's Republic of China.
authors.forename: J W
authors.initials: JW
authors.lastname: Wu
authors.forename: Z X
authors.initials: ZX
authors.lastname: Wang
authors.forename: J M
authors.initials: JM
authors.lastname: Zhou
mesh_terms: D000818:Animals; D006224:Cricetinae; D003412:Cricetulus; D004789:Enzyme Activation; D019791:Guanidine / Q000378:metabolism*; D007700:Kinetics; D017510:Protein Folding; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry / Q000378:metabolism*
chemical_list: D013762:Tetrahydrofolate Dehydrogenase; D019791:Guanidine