Acarbose binding at the surface of Saccharomycopsis fibuligera glucoamylase suggests the presence of a raw starch-binding site

There are two acarbose molecules in the structure of the complex of glucoamylase from Saccharomycopsis fibuligera HUT 7212 (Glu) with the inhibitor acarbose: one is bound to the active site, which is localized in the cavity of the (alpha/alpha)(6)-barrel, the other one is on the surface of the enzyme molecule curved wound Tyr464, which captures the inhibitor molecule as seen in the "sugar tongs" binding site of the barley alpha-amylase isozyme I complexed with a thiomaltooligosaccharide. The surface acarbose molecule suggests the presence of a raw starch-binding site. Based on the expected similarity of structures of glucoamylase Glu, which does not degrade raw starch, and the raw starch-degrading glucoamylase from S. fibuligera IFO 0111 (Glm), it is reasonable to expect the presence of a starch-binding site, at a structurally equivalent position, also on the surface of Glm. Verification of this hypothesis by preparation of mutants of glucoamylases Glu and Glm is presented.