ABSTRACT
The study of the kinetics of thermal aggregation of glycogen phosphorylase b (Phb) from rabbit skeletal muscles by dynamic light scattering at 48°C showed that 2-hydroxypropyl-ß-cyclodextrin (HP-ß-CD) accelerated the aggregation process and induced the formation of the larger protein aggregates. The reason of the accelerating effect of HP-ß-CD is destabilization of the protein molecule under action of HP-ß-CD. This conclusion was supported by the data on differential scanning calorimetry and the kinetic data on thermal inactivation of Phb. It is assumed that destabilization of the Phb molecule is due to preferential binding of HP-ß-CD to intermediates of protein unfolding in comparison with the original native state. The conclusion regarding the ability of the native Phb for binding of HP-ß-CD was substantiated by the data on the enzyme inhibition by HP-ß-CD. © 2010 Wiley Periodicals, Inc. Biopolymers 93: 986-993, 2010.
Subject(s)
Glycogen Phosphorylase, Muscle Form/chemistry , Glycogen Phosphorylase, Muscle Form/drug effects , beta-Cyclodextrins/pharmacology , 2-Hydroxypropyl-beta-cyclodextrin , Animals , Enzyme Stability/drug effects , Glycogen Phosphorylase, Muscle Form/metabolism , In Vitro Techniques , Kinetics , Light , Muscle, Skeletal/enzymology , Protein Multimerization/drug effects , Rabbits , Scattering, Radiation , ThermodynamicsABSTRACT
Kinetic analysis of the glycogen chain growth reaction catalyzed by glycogen phosphorylase b from rabbit skeletal muscle has been carried out over a wide range of AMP concentration under the saturation of the enzyme by glycogen. Applicability of some variants of the kinetic model involving the interaction of AMP- and glucose 1-phosphate-binding sites in the dimeric enzyme molecule is considered. A kinetic model of the enzymatic reaction describing adequately the activation of the enzyme by AMP and inhibition at sufficiently high concentrations of AMP is proposed.
