Isolation and overexpression of a gene encoding an extracellular beta-(1,3-1,4)-glucanase from Streptococcus bovis JB1

Appl Environ Microbiol. 1997 Oct;63(10):3752-6. doi: 10.1128/aem.63.10.3752-3756.1997.

Abstract

Streptococcus bovis JB1 was found to produce a 25-kDa extracellular enzyme active against beta-(1,3-1,4)-glucans. A gene was isolated encoding a specific beta-(1,3-1,4)-glucanase that corresponds to this size and belongs to glycoside hydrolase family 16. A 4- to 10-fold increase in supernatant beta-glucanase activity was obtained when the cloned beta-glucanase gene was reintroduced into S. bovis JB1 by use of constructs based on the plasmid vector pTRW10 or pIL253. The beta-(1,3-1,4)-glucanase gene was also expressed upon introduction of the pTRW10 construct pTRWL1R into Lactococcus lactis IL2661 and Enterococcus faecalis JH2-SS, although extracellular activity was 8- to 50-fold lower than that in S. bovis JB1. The beta-(1,3-1,4)-glucanase purified from the culture supernatant of S. bovis JB1 carrying pTRWL1R showed a K(m) of 2.8 mg per ml and a Vmax of 338 mumol of glucose equivalents per min per mg of protein with barley beta-glucan as the substrate. The S. bovis beta-(1,3-1,4)-glucanase may contribute to the ability of this bacterium to utilize starch by degrading structural polysaccharides present in endosperm cell walls.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • Cloning, Molecular
  • DNA, Bacterial / genetics
  • Endo-1,3(4)-beta-Glucanase*
  • Enterococcus faecalis / genetics
  • Extracellular Space / enzymology
  • Gene Expression
  • Genes, Bacterial*
  • Genetic Vectors
  • Glycoside Hydrolases / genetics*
  • Glycoside Hydrolases / isolation & purification
  • Glycoside Hydrolases / metabolism
  • Lactococcus lactis / genetics
  • Molecular Sequence Data
  • Plasmids / genetics
  • Recombinant Proteins / genetics
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Streptococcus bovis / enzymology*
  • Streptococcus bovis / genetics*

Substances

  • DNA, Bacterial
  • Recombinant Proteins
  • Glycoside Hydrolases
  • Endo-1,3(4)-beta-Glucanase