Subsite Structure of Saccharomycopsis α-Amylase Secreted from Saccharomyces cerevisiae

Abstract

The kinetic parameters (k_cat/K_m) and the cleaved-bond distributions for the hydrolysis of linear maltooligosaccharides G_n(3λn λ9) by Saccharomycopsis α-amylase (Sfamy) secreted from Saccharomyces cerevisiae were determined at pH5.25 and 25°C. The subsite affinities of Sfamy were also evaluated from these data. The subsite structure of Sfamy is characteristic of the active site of an endo-cleavage type enzyme, consisting of internal repulsive sites with the catalytic residues and external attractive sites. Moreover, the pK_a, values of the catalytic residues were calculated from the pH dependence plot of the kinetic parameter (k_cat/K_m). The amino acid residues which contribute to the subsite affinities and the catalytic activity of Sfamy are proposed and compared with those of Taka-amylase A.