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The composition and the structures of native “group B saponin” in soybean seeds were reinvestigated. Five kinds of saponins named soyasaponins αg, βg, βa, γg, and γa, according to elution order from HPLC, were isolated and the structures were characterized as 2,3-dihydro-2,5-dihydroxy-6-methyl-4H-pyran-4-one (DDMP) attaching through an acetal linkage to the C-22 hydroxyl of the aglycones of soyasaponins V, I, II, III, and IV, respectively, by UV, IR, MS, and NMR. DDMP-conjugated saponins were detected as major saponin constituents by extraction under mild conditions, and soyasaponins I–V were not detected. Therefore it was strongly suggested that these DDMP-conjugated saponins were genuine saponins in the intact soybeans.

alpha-Glucosidase was partially purified 103-fold from a cell-free extract of Torulaspora pretoriensis YK-1 by column chromatography on Toyopearl HW55F, DEAE-Toyopearl 650M, hydroxylapatite and phenyl-Toyopearl 650M. Further purification by preparative polyacrylamide gel electrophoresis (PAGE) gave the homogenous protein, but the specific activity was reduced. The molecular weight of the enzyme was estimated to be 69,000 by SDS-PAGE and 60,000 by gel filtration. Optimum pH and temperature were 6.8 and 35 degrees C, respectively. The enzyme was inhibited strongly by AgNO3, HgCl2, sodium dodecyl sulfate, and N-ethylmaleimide. The Km (mM) for p-nitrophenyl alpha-D-glucopyranoside, maltose, maltotriose, isomaltose, methyl alpha-glucoside, and sucrose were 0.15, 150, 45, 17, 18, and 29, and Vmax (mumol/min/mg protein) for those substrates were 87, 0.23, 2.4, 9.0, 12, and 7.4, respectively. The N-terminal amino acid sequence of the enzyme was PEVKNHPETQPKWWKEATVY. The properties of alpha-glucosidase from T. pretoriensis YK-1 were similar to those from Saccharomyces cerevisiae.

γ-Glutamylmethylamide synthetase [L-glutamate: methylamine ligase (ADP-forming), EC 6.3.4.12] was purified about 70-fold from a cell-free extract of Methylophaga sp. AA-30 by ammonium sulfate fractionation, Octyl-Sepharose column chromatography, and Sephacryl S-300 gel filtration. Only a single protein band was detected after SDS-polyacrylamide gel electrophoresis of the purified preparation; the band was at a position corresponding to a molecular weight of 56,000. The molecular weight of the enzyme was calculated to be 440,000 by Superose 6HR gel filtration, so we suggest that the enzyme is an octomer of identical subunits. The enzyme had maximum activity at pH 7.5 and 40°C. It could use ethylamine and propylamine instead of methylamine as the substrate, but it could not use D-glutamate or L-glutamine instead of L-glutamate.

A glass microfiber filter was found useful for detection of the complex of lysyl-tRNA synthetase from Bacillus stearothermophilus with lysyladenylate. Detected c.p.m. values of the 3H-labeled complex trapped by the glass microfiber filter was about 2-fold higher than those by nitrocellulose filter and about 3-fold higher than those by DEAE-cellulose filter because of its higher counting efficiency. The use of the glass fiber filter is more suitable for the lysyl-tRNA synthetase assay than the use of either nitrocellulose or DEAE-cellulose filters, which were used for other aminoacyl-tRNA synthetases. Effects of pH on the filter assay of lysyl-tRNA synthetase activity using the glass microfiber filter were also examined.

Invertase was purified from Torulaspora pretoriensis YK-1 by acid treatment and column chromatography on DEAE-Toyopearl 650M and phenyl-Toyopearl 650M to homogeneity. The molecular weight of the purified enzyme was estimated to be 130,000 by SDS-polyacrylamide gel electrophoresis and 530,000 by gel filtration. The enzyme contained 50% molecular weight as carbohydrate. Properties of the invertase from T. pretoriensis was similar to external invertase from Saccharomyces cerevisiae.

Among the yeasts belonging to the genus Torulaspora, T. pretoriensis IFO 10218 was selected as a maltose-fermenting strain that could leaven dough containing 5% glucose as high as commercial baking strains. The freeze-thaw resistance of this strain in the dough was higher than that of Saccharomyces cerevisiae FL 2209 isolated from commercial compressed yeast. Since the cells of IFO 10218 readily sedimented in their suspension, a dispersing mutant, YK-1, was induced. When YK-1 was cultured in the. presence of NaCl, leavening abilities were reduced, unlike FL 2209. α-Glucosidase activity of YK-1 cells grown on maltose increased slightly, resulting in partly improved leavening ability in the dough without addition of sugar. YK-1 was shown to be a candidate applicable to breadmaking by not only usual method but also the frozen-dough method.

(1992). Purification, Crystallization, and Properties of the Extracellular Levansucrase from Zymomonas mobilis. Bioscience, Biotechnology, and Biochemistry: Vol. 56, No. 8, pp. 1335-1337.

(1992). Isolation and Structural Elucidation of the Major Genuine Soybean Saponin. Bioscience, Biotechnology, and Biochemistry: Vol. 56, No. 1, pp. 142-143.