Influence of Dielectric Constants and Ligand Binding on Thermostability of Glucoamylase

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Abstract
Thermal stability of glucoamylase [α-1,4: 1,6-glucan-4: 6-glucohydrolase, EC 3.2.1.3] from Rhizopus niveus was investigated in detail mainly at 60.0±0.1°C (pH 4.5) in relation to kinetics both in the presence and in the absence of various ligands. With substrate analogues, such as glucose, lactose and gluconolactone, and with glycerol, thermostability of the enzyme greatly increased, whereas decreased with dioxane, ethanol and glycol. To elucidate such phenomena clearly, several model equations were propounded on the assumption that there were two main factors which would play an essential role in heat stability of the enzyme: binding of ligands and the dielectric constants of solvents. From the model equations, binding constants of the ligands were estimated in order to confirm the validity of the assumption; e.g., the inhibitor constant for glucose (at 59.5°C) corresponding to the reciprocal of binding constant was evaluated to be 0.25 m as was expected.