Thermodynamic characterization of hog kidney D-amino acid oxidase apoenzyme in concentrated guanidine hydrochloride solution. Preferential interaction with the solvent components and the molecular weight of the monomeric unit.
This paper describes the physical characterization of the monomeric unit of hog kidney D-amino acid oxidase apoenzyme in 6 M guanidine hydrochloride (GuHCl) solution by means of differential refractometry, densimetry, light scattering, equilibrium sedimentation, and high-speed gel filtration chromatography. In 6 M GuHCl solution, the oxidase interacts preferentially with GuHCl: the values of the preferential interaction parameter are 0.11 +/- 0.03 (S.D.) g/g of protein by densimetry and 0.14 +/- 0.04 g/g of protein by refractometry. The volume change, delta V, of the oxidase on transfer from the native to the denatured state is -350 ml/mol. The molecular weight of the monomeric apoenzyme is 39,600 +/- 1,700 by light scattering and 38,000 +/- 1,200 by high-speed equilibrium sedimentation. The values of the molecular weight estimated by the empirical methods, i.e., sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis and high-speed gel filtration chromatography in 6 M GuHCl, agree well with those obtained by the thermodynamic methods mentioned above. These results confirm definitely that the complex of the apoenzyme with SDS normally behaves in the same manner as those of standard proteins in SDS-gel electrophoresis. This is also supported in this study by the analysis of the electrophoretic data at several gel concentrations by Ferguson plots. The molecular weight of quasi-D-amino acid oxidase apoenzyme was also examined by the empirical methods.
D000818:Animals; D001051:Apoenzymes / Q000032:analysis; D055598:Chemical Phenomena; D002621:Chemistry; D002850:Chromatography, Gel; D003605:D-Amino-Acid Oxidase / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel; D006146:Guanidines; D007668:Kidney / Q000201:enzymology*; D008970:Molecular Weight; D013552:Swine; D013816:Thermodynamics
D001051:Apoenzymes; D006146:Guanidines; D003605:D-Amino-Acid Oxidase