Substrate specificity of solvent viscosity effects in carboxypeptidase A catalyzed peptide hydrolysis.
We have investigated the viscosity of carboxypeptidase A catalyzed Bz-Gly-Phe hydrolysis at pH 7.5 (Tris) and 0.5 mol.l-1 NaCl over the range 10-100 mp, varied by addition of glycerol or sucrose. In contrast to previous reports of strong viscosity effects on the corresponding Cbz-Ala-Ala-Ala hydrolysis, both the catalytic constant and the Michaelis constant are virtually independent of viscosity over the 10-fold range investigated. Furthermore, the CD spectra of carboxypeptidase A in the high-viscosity media point to no change in the alpha-helix and beta-sheet structure in these media. The data are compatible either with a compacter, more rigid enzyme-substrate structure or with a more prominent role of intramolecular nuclear reorganization compared to protein reorganization for Bz-Gly-Phe than for Cbz-Ala-Ala-Ala. These views can be given a preciser frame in terms of stochastic chemical rate theory.
Khoshtariya
D E
DE
Institute of Inorganic Chemistry andElectrochemistry, Georgian Academy of Sciences, Tbilisi, U.S.S.R.
Hammerstad-Pedersen
J M
JM
Ulstrup
J
J
D000595:Amino Acid Sequence; D000818:Animals; D002268:Carboxypeptidases / Q000378:metabolism*; D043422:Carboxypeptidases A; D002417:Cattle; D002942:Circular Dichroism; D007700:Kinetics; D008433:Mathematics; D008969:Molecular Sequence Data; D009842:Oligopeptides; D010179:Pancreas / Q000201:enzymology; D011487:Protein Conformation; D012997:Solvents; D013379:Substrate Specificity; D014783:Viscosity
D009842:Oligopeptides; D012997:Solvents; D002268:Carboxypeptidases; D043422:Carboxypeptidases A