Alteration of the properties of Aspergillus sp. K-27 glucoamylase on limited proteolysis with subtilisin.

An active derivative (mol. wt. 48,000) of Aspergillus sp. K-27 glucoamylase (mol. wt. 76,000) was obtained by limited proteolysis with subtilisin. The amino acid sequences of native and modified enzymes at the N-termini were Ala-Gly-Gly-Thr-Leu-Asp and Ala-Val-Leu, respectively. The proteolysis greatly decreased the affinity of the enzyme for amylopectin and glycogen, but not for oligosaccharides. It also reduced the ability of the enzyme to degrade raw starch, abolished the ability of the enzyme to adsorb onto starch granules, and eliminated the synergistic action of the enzyme in the hydrolysis of starch granules with alpha-amylase. These findings imply that the enzyme has a specific affinity site for polysaccharide substrates besides the catalytic site, i.e., a starch-binding site, and that the former is removed by proteolysis. The extent of the reduction in the activity for raw starches caused by the modification varied with the starch source, as the modified enzyme digested raw potato starch better than either raw corn or sweet potato starches. A new method for evaluation of the raw starch-digesting activity of glucoamylase is described.