Purification, characterization, and overexpression of an endo-1,4-β-mannanase from thermotolerant Bacillus sp. SWU60

World J Microbiol Biotechnol. 2017 Mar;33(3):53. doi: 10.1007/s11274-017-2224-7. Epub 2017 Feb 20.

Abstract

Endo-β-1,4-mannanases are important catalytic agents in several industries. The enzymes randomly cleave the β-1,4-linkage in the mannan backbone and release short β-1,4-mannooligosaccharides and mannose. In the present study, mannanase (ManS2) from thermotolerant Bacillus sp. SWU60 was purified, characterized, and its gene was cloned and overexpressed in Escherichia coli. ManS2 was purified from culture filtrate (300 ml) by using hydrophobic, ion-exchange, and size-exclusive liquid chromatography. The apparent molecular mass was 38 kDa. Optimal pH and temperature for enzyme activity were 6.0 and 60 °C, respectively. The enzyme was stable up to 60 °C for 1 h and at pH 5-9 at 4 °C for 16 h. Its enzyme activity was inhibited by Hg2+. The full-length mans2 gene was 1,008 bp, encoding a protein of 336 amino acids. Amino acid sequence analysis revealed that it belonged to glycoside hydrolase family 26. Konjac glucomannan was a favorable substrate for recombinant ManS2 (rManS2). rManS2 also degraded galactomannan from locust bean gum, indicating its potential for production of glucomanno- and galactomanno-oligosaccharides. Both native and recombinant ManS2 from Bacillus sp. SWU60 can be applied in several industries especially food and feed.

Keywords: Bacillus sp.; Endo-β-1,4-mannanases; Galactomannan; Glucomannan; Mannan; Recombinant enzyme.

MeSH terms

  • Bacillus / chemistry
  • Bacillus / enzymology*
  • Bacillus / genetics
  • Bacillus / isolation & purification
  • Base Sequence
  • Chromatography, Ion Exchange / methods
  • Enzyme Activation
  • Enzyme Stability
  • Escherichia coli / genetics
  • Mannans / metabolism
  • Substrate Specificity
  • beta-Mannosidase / biosynthesis*
  • beta-Mannosidase / chemistry*
  • beta-Mannosidase / genetics
  • beta-Mannosidase / isolation & purification

Substances

  • Mannans
  • galactomannan
  • beta-Mannosidase