Enzymatic and genetic characterization of the DasD protein possessing N-acetyl-β-d-glucosaminidase activity in Streptomyces coelicolor A3(2)

FEMS Microbiol Lett. 2013 Mar;340(1):33-40. doi: 10.1111/1574-6968.12069. Epub 2013 Jan 16.

Abstract

The dasD gene is located just downstream of the dasABC gene cluster, encoding components of an ABC transporter for uptake of a chitin-degradation product N,N'-diacetylchitobiose [(GlcNAc)(2) ] in Streptomyces coelicolor A3(2). To clarify the roles of the DasD protein in the degradation and assimilation of chitin, we obtained and characterized a recombinant DasD protein and a dasD-null mutant of S. coelicolor A3(2). The recombinant DasD protein produced in Escherichia coli showed N-acetyl-β-d-glucosaminidase (GlcNAcase) activity and its optimum temperature and pH were 40 °C and 7, respectively. dasD transcription was strongly induced in the presence of chitin, weakly by chitosan, but not by cellulose or xylan in S. coelicolor A3(2). Immuno-blot analysis demonstrated that DasD is a cytoplasmic protein. The dasD-null mutant exhibited cellular GlcNAcase activity which was comparable with that of the parent strain M145. DasD, thus, did not seem to be a major GlcNAcase. Induced extracellular chitinase activity in the dasD-null mutant was, interestingly, higher than M145, in the presence of colloidal chitin or (GlcNAc)(2) . In contrast to M145, (GlcNAc)(2) temporally accumulated in the culture supernatant of the dasD-null mutant in the presence of colloidal chitin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetylglucosaminidase / genetics*
  • Acetylglucosaminidase / metabolism*
  • Chitin / metabolism
  • Cloning, Molecular
  • Cytoplasm / enzymology
  • Enzyme Stability
  • Escherichia coli
  • Gene Deletion
  • Gene Expression Regulation, Bacterial
  • Gene Expression Regulation, Enzymologic
  • Hydrogen-Ion Concentration
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Streptomyces coelicolor / enzymology*
  • Streptomyces coelicolor / genetics
  • Streptomyces coelicolor / metabolism
  • Temperature

Substances

  • Recombinant Proteins
  • Chitin
  • Acetylglucosaminidase