Furostanol glycoside 26-O-beta-glucosidase from the leaves of Solanum torvum.
A beta-glucosidase (torvosidase) was purified to homogeneity from the young leaves of Solanum torvum. The enzyme was highly specific for cleavage of the glucose unit attached to the C-26 hydroxyl of furostanol glycosides from the same plant, namely torvosides A and H. Purified torvosidase is a monomeric glycoprotein, with a native molecular weight of 87 kDa by gel filtration and a pI of 8.8 by native agarose IEF. Optimum pH of the enzyme for p-nitrophenyl-beta-glucoside and torvoside H was 5.0. Kinetic studies showed that Km values for torvoside A (0.06 3mM) and torvoside H (0.068 mM) were much lower than those for synthetic substrates, pNP-beta-glucoside (1.03 mM) and 4-methylumbelliferyl-beta-glucoside (0.78 mM). The enzyme showed strict specificity for the beta-d-glucosyl bond when tested for glycone specificity. Torvosidase hydrolyses only torvosides and dalcochinin-8'-beta-glucoside, which is the natural substrate of Thai rosewood beta-glucosidase, but does not hydrolyse other natural substrates of the GH1 beta-glucosidases or of the GH3 beta-glucosidase families. Torvosidase also hydrolyses C5-C10 alkyl-beta-glucosides, with a rate of hydrolysis increasing with longer alkyl chain length. The internal peptide sequence of Solanum beta-glucosidase shows high similarity to the sequences of family GH3 glycosyl hydrolases.
Arthan
Dumrongkiet
D
Department of Biochemistry and Center for Excellence in Protein Structure and Function, Faculty of Science, Mahidol University, Rama VI Road, Bangkok 10400, Thailand.
Kittakoop
Prasat
P
Esen
Asim
A
Svasti
Jisnuson
J
D000595:Amino Acid Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D006027:Glycosides / Q000138:chemical synthesis / Q000737:chemistry; D006868:Hydrolysis; D007700:Kinetics; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D018515:Plant Leaves / Q000737:chemistry / Q000201:enzymology*; D012503:Saponins / Q000138:chemical synthesis / Q000737:chemistry; D016415:Sequence Alignment; D032322:Solanum / Q000201:enzymology*; D001617:beta-Glucosidase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification
D006027:Glycosides; D012503:Saponins; C454584:torvoside A; C509529:torvoside H; C097713:furostanol glycoside 26-O-beta-glucosidase; D001617:beta-Glucosidase