Furostanol glycoside 26-O-β-glucosidase from the leaves of Solanum torvum

Abstract

A β-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-β-glucoside and torvoside H was 5.0. Kinetic studies showed that Kmvalues for torvoside A (0.063 mM) and torvoside H (0.068 mM) were much lower than those for synthetic substrates, pNP-β-glucoside (1.03 mM) and 4-methylumbelliferyl-β-glucoside (0.78 mM). The enzyme showed strict specificity for the β-d-glucosyl bond when tested for glycone specificity. Torvosidase hydrolyses only torvosides and dalcochinin-8′- β-glucoside, which is the natural substrate of Thai rosewood β-glucosidase, but does not hydrolyse other natural substrates of the GH1 β-glucosidases or of the GH3 β-glucosidase families. Torvosidase also hydrolyses C5-C10alkyl-β-glucosides, with a rate of hydrolysis increasing with longer alkyl chain length. The internal peptide sequence of Solanum β-glucosidase shows high similarity to the sequences of family GH3 glycosyl hydrolases. © 2005 Elsevier Ltd. All rights reserved.