title: Two slow stages in refolding of bovine carbonic anhydrase B are due to proline isomerization.
abstract: Kinetics of refolding of bovine carbonic anhydrase B have been studied by the "double-jump" technique (i.e. the dependence of protein refolding on delay time in the unfolded state after fast unfolding). It is shown that two stages (the slow with a relaxation time of t1/2 approximately equal to 120 s and the superslow with t1/2 approximately equal to 600 s) observed during refolding of bovine carbonic anhydrase B are due to trans-cis isomerization of proline residues. The dependences of rate constants of these processes on temperature and on the final denaturant concentration were measured. Activation energies of both processes are the same, Ea = 18(+/- 2) kcal/mol. The rate constants of protein refolding do not depend on the final concentration of urea under native conditions. In addition, the rate of isomerization of essential proline residues in the "molten globule" intermediate state of bovine carbonic anhydrase was measured and found to be equal to that for unstructural polypeptides. The effect of several proline residues on carbonic anhydrase refolding is discussed.
authors.affiliation: Institute of Protein Research, Academy of Sciences of the U.S.S.R. Pushchino, Moscow Region.
authors.forename: G V
authors.initials: GV
authors.lastname: Semisotnov
authors.forename: V N
authors.initials: VN
authors.lastname: Uversky
authors.forename: I V
authors.initials: IV
authors.lastname: Sokolovsky
authors.forename: A M
authors.initials: AM
authors.lastname: Gutin
authors.forename: O I
authors.initials: OI
authors.lastname: Razgulyaev
authors.forename: N A
authors.initials: NA
authors.lastname: Rodionova
mesh_terms: D000818:Animals; D002256:Carbonic Anhydrases; D002417:Cattle; D002942:Circular Dichroism; D004912:Erythrocytes / Q000201:enzymology*; D007536:Isomerism; D007700:Kinetics; D011392:Proline / Q000378:metabolism*; D011487:Protein Conformation; D011489:Protein Denaturation; D013696:Temperature
chemical_list: D011392:Proline; D002256:Carbonic Anhydrases