Screening of a xylanase clone from a fosmid library of rumen microbiota in Hu sheep

Anim Biotechnol. 2012;23(3):156-73. doi: 10.1080/10495398.2012.662925.

Abstract

The glycosyl hydrolase family 11, which is responsible for carbohydrate metabolism, was identified in the open reading frame (ORF) 6 of a xylanase positive clone from a fosmid library of rumen microbiota of Hu sheep. A BLASTP search of GenBank revealed that ORF6 encoded a 355-amino acid putative endoxylanase, having 61% similarity (e(-73)) to endo-1,4-β-xylanase of Fibrobacter succinogenes S85 (YP_003250510.1). Predicted with the SWISS-MODEL, there were two separate β-sandwich clusters linked with a high serine containing linker in ORF6. The N-terminal β-sandwich is a novel endoxylanase of the glycosyl hydrolase family 11 with a specific activity of 1150.00 U/mg. The optimal pH and temperature for this enzyme were shown to be pH 5.0 and 50°C, respectively. The C-terminal helped increase the stability of the xylanase but decreased the activity to some degree. The C-terminal β-sandwich could bind avicel, but no conserved domain could be found. It may be a novel carbohydrate-binding module.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics*
  • Bacterial Proteins / metabolism
  • Base Sequence
  • Biotechnology
  • Cellulose / metabolism
  • Cloning, Molecular
  • DNA Primers / genetics
  • Endo-1,4-beta Xylanases / chemistry
  • Endo-1,4-beta Xylanases / genetics*
  • Endo-1,4-beta Xylanases / metabolism
  • Enzyme Stability
  • Fibrobacter / enzymology
  • Fibrobacter / genetics
  • Hydrogen-Ion Concentration
  • Kinetics
  • Metagenome
  • Molecular Sequence Data
  • Molecular Weight
  • Open Reading Frames
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Rumen / microbiology*
  • Sequence Homology, Amino Acid
  • Sheep / microbiology*
  • Temperature

Substances

  • Bacterial Proteins
  • DNA Primers
  • Recombinant Proteins
  • Cellulose
  • Endo-1,4-beta Xylanases