Substrate Specificity and Subsite Affinities of Crystalline α-Glucosidase from Aspergillus niger(Biological Chemistry)

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Asp. niger α-glucosidase was crystallized in ammonium sulfate solution after chromatographies on DEAE-Sepharose CL-6B and TOYOPEARL HW-55 columns. The crystalline α-glucosidase, which was a glycoprotein containing 25.5% carbohydrate as glucose, gave a single band on polyacrylamide disc gel electrophoresis. The molecular weight was estimated to be about 12.5×10^4 by SDS-disc gel electrophoresis. However, the crystalline enzyme consists of two components (M.W. about 3.3×10^4 and 9.8×10^4, respectively) separable only by reverse-phase HPLC causing irreversible inactivation. The optimum pH was 4.3. The enzyme also hydrolyzed α-glucans such as soluble starch and amylose. The ratios of V values (Km values, in parentheses, mM of nonreducing terminal) for maltose, kojibiose, nigerose, isomaltose, phenyl α-glucoside, phenyl α-maltoside, malto-triose, -tetraose, -oentaose, -hexaose, -heptaose and -octaose, maltodextrin (DP=17), and soluble starch were estimated to be 100 (0.75) : 23 (4.6) : 62 (12) : 35 (8.0) : 1.3 (0.34) : 126 (0.87) : 126 (0.69) : 135 (1.1) : 102 (1.9) : 119 (3.2) : 102 (4.9) : 89 (5.3) : 89 (11) : 114 (4.3). The subsite affinities in the active site were 0.790, 5.93 and 0.191 kcal/mol for subsites 1, 2, and 3, respectively. The three subsites were considered to be effective for the binding of substrate.
社団法人日本農芸化学会の論文
1991-09-23