Shuzo Sakai

The crystal structure and physicochemical properties of l-ascorbic acid 2-glucoside

The stable L-ascorbic acid glucoside produced by the action of the cyclomaltodextrin glucanotransferase (CGTase, EC 2.4.1.19) from Bacillus stearothermophilus was crystallized from an aqueous solution. Determination of the molecular structure by single crystal X-ray analysis showed the compound to be 2-O-alpha-D-glucopyranosyl-L-ascorbic acid (AA-2G). The crystals are orthorhombic, space group P2(1)2(1)2(1), with unit-cell dimensions a = 11.929 A, b = 24.351 A, and c = 4.864 A. The D-glucopyranose residue has the 4C1 conformation. These conclusions are in good agreement with those based on the 13C-NMR spectrum. The general physicochemical properties of crystalline AA-2G are reported.

Crystalline D-mannitol:NAD+ oxidoreductase from Leuconostoc mesenteroides

The crystalline D-mannitol dehyrogenase (D-mannitol:NAD oxidoreductase, EC 1.1.1.67) catalyzed the reversible reduction of D-fructose to D-mannitol. D-Sorbitol was oxidized only at the rate of 40 of the activity for D-mannitol. The enzyme was inactive for all of four pentitols and their corresponding 2-ketopentoses. The apparent optimal pH for the reduction of D-fructose or the oxidation of D-mannitol was 5.35 or 8.6, respectively. The Michaelis constants were 0.035M for D-fructose and 0.020M for D-mannitol. The enzyme was also found to be specific for NAD. The Michaelis constans were 1 x 10-5M for NADH2 and 2.7 x 10-4M for NAD.

Crystalline d-Mannitol:NAD Oxidoreductase from Leuconostoc mesenteroides: Part II. Substrate and Coenzyme Specificity

The crystalline d-mannitol dehyrogenase (d-mannitol:NAD oxidoreductase, EC 1.1.1.67) catalyzed the reversible reduction of d-fructose to d-mannitol. d-Sorbitol was oxidized only at the rate of 4% of the activity for d-mannitol. The enzyme was inactive for all of four pentitols and their corresponding 2-ketopentoses. The apparent optimal pH for the reduction of d-fructose or the oxidation of d-mannitol was 5.35 or 8.6, respectively. The Michaelis constants were 0.035 m for d-fructose and 0.020 m for d-mannitol. The enzyme was also found to be specific for NAD. The Michaelis constans were 1 × 10−5 m for NADH2 and 2.7 × 10−4 m for NAD.