Intra- and extracellular beta-galactosidases from Bifidobacterium bifidum and B. infantis: molecular cloning, heterologous expression, and comparative characterization

Appl Environ Microbiol. 2001 May;67(5):2276-83. doi: 10.1128/AEM.67.5.2276-2283.2001.

Abstract

Three beta-galactosidase genes from Bifidobacterium bifidum DSM20215 and one beta-galactosidase gene from Bifidobacterium infantis DSM20088 were isolated and characterized. The three B. bifidum beta-galactosidases exhibited a low degree of amino acid sequence similarity to each other and to previously published beta-galactosidases classified as family 2 glycosyl hydrolases. Likewise, the B. infantis beta-galactosidase was distantly related to enzymes classified as family 42 glycosyl hydrolases. One of the enzymes from B. bifidum, termed BIF3, is most probably an extracellular enzyme, since it contained a signal sequence which was cleaved off during heterologous expression of the enzyme in Escherichia coli. Other exceptional features of the BIF3 beta-galactosidase were (i) the monomeric structure of the active enzyme, comprising 1,752 amino acid residues (188 kDa) and (ii) the molecular organization into an N-terminal beta-galactosidase domain and a C-terminal galactose binding domain. The other two B. bifidum beta-galactosidases and the enzyme from B. infantis were multimeric, intracellular enzymes with molecular masses similar to typical family 2 and family 42 glycosyl hydrolases, respectively. Despite the differences in size, molecular composition, and amino acid sequence, all four beta-galactosidases were highly specific for hydrolysis of beta-D-galactosidic linkages, and all four enzymes were able to transgalactosylate with lactose as a substrate.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Bifidobacterium / classification
  • Bifidobacterium / enzymology*
  • Bifidobacterium / genetics
  • Binding Sites / genetics
  • Escherichia coli / enzymology
  • Escherichia coli / genetics
  • Genes, Bacterial
  • Molecular Sequence Data
  • Recombinant Proteins / metabolism
  • Sequence Analysis, DNA
  • Substrate Specificity
  • beta-Galactosidase* / chemistry
  • beta-Galactosidase* / genetics
  • beta-Galactosidase* / isolation & purification
  • beta-Galactosidase* / metabolism

Substances

  • Recombinant Proteins
  • beta-Galactosidase

Associated data

  • GENBANK/AJ224434
  • GENBANK/AJ224435
  • GENBANK/AJ224436
  • GENBANK/AJ272131