Influence of Molecular Structures of Substrates and Analogues on Taka-amylase A Catalyzed HydrolysesI. Effect of Chain Length of Linear Substrates*

Abstract

1) Hydrolyses of linear maltodextrins (DP 2–DPn 117) catalyzed by crystalline Takaamylase A [EC 3. 2.1.1] were studied at pH 5. 3 and 25°C to determine the rate parameter for each substrate. 2) Michaelis constant, Km (in moler basis), decreased with increasing chain length (n]of glucose unit. Turnover number, V/(E)0, increased with chain length up to n= 7 and became practically constant for n more than 7, which suggests that the specificity region of Taka-amylase A spans about 7 glucose units. 3) The dependence of Km and V/(E)0 on chain length of substrate was interpreted in terms of the probability of formation of productive and nonproductive ES-complexes. Based on the assumption that the breakdown rate constant in productive ES-complex, ktnt, is constant irrespective of chain length of substrate and binding mode, magnitudes of subsite affinities (in free energy units) in the active site were evaluated from the values of V/Km(E)0for various substrates. Results suggested the presence of a distortion in the productive ES complex.