Characterization of a chimeric enzyme comprising feruloyl esterase and family 42 carbohydrate-binding module

Appl Microbiol Biotechnol. 2010 Mar;86(1):155-61. doi: 10.1007/s00253-009-2224-0. Epub 2009 Sep 16.

Abstract

We engineered a chimeric enzyme (AwFaeA-CBM42) comprising of type-A feruloyl esterase from Aspergillus awamori (AwFaeA) and family 42 carbohydrate-binding module (AkCBM42) from glycoside hydrolase family 54 alpha-L-arabinofuranosidase of Aspergillus kawachii. The chimeric enzyme was successfully produced in Pichia pastoris and accumulated in the culture broth. The purified chimeric enzyme had an apparent relative molecular mass (M(r)) of 53,000. The chimeric enzyme binds to arabinoxylan; this indicates that the AkCBM42 in AwFaeA-CBM42 binds to arabinofuranose side chain moiety of arabinoxylan. The thermostability of the chimeric enzyme was greater than that of AwFaeA. No significant difference of the specific activity toward methyl ferulate was observed between the AwFaeA and chimeric enzyme, but the release of ferulic acid from insoluble arabinoxylan by the chimeric enzyme was approximately 4-fold higher than that achieved by AwFaeA alone. In addition, the chimeric enzyme and xylanase acted synergistically for the degradation of arabinoxylan. In conclusion, the findings of our study demonstrated that the components of the AwFaeA-CBM42 chimeric enzyme act synergistically to bring about the degradation of complex substrates and that the family 42 carbohydrate-binding module has potential for application in the degradation of polysaccharides.

Publication types

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

MeSH terms

  • Arabinose / analogs & derivatives
  • Arabinose / metabolism
  • Aspergillus / enzymology*
  • Aspergillus / genetics
  • Aspergillus / metabolism
  • Biotechnology / methods*
  • Carbohydrate Metabolism
  • Carboxylic Ester Hydrolases* / chemistry
  • Carboxylic Ester Hydrolases* / genetics
  • Carboxylic Ester Hydrolases* / metabolism
  • Coumaric Acids / metabolism
  • Fungal Proteins / chemistry
  • Fungal Proteins / genetics
  • Fungal Proteins / metabolism
  • Glycoside Hydrolases* / chemistry
  • Glycoside Hydrolases* / genetics
  • Glycoside Hydrolases* / metabolism
  • Pichia / enzymology
  • Pichia / genetics
  • Protein Engineering / methods
  • Recombinant Fusion Proteins* / chemistry
  • Recombinant Fusion Proteins* / genetics
  • Recombinant Fusion Proteins* / isolation & purification
  • Recombinant Fusion Proteins* / metabolism
  • Substrate Specificity
  • Xylans / metabolism

Substances

  • Coumaric Acids
  • Fungal Proteins
  • Recombinant Fusion Proteins
  • Xylans
  • arabinofuranose
  • arabinoxylan
  • ferulic acid
  • Arabinose
  • Carboxylic Ester Hydrolases
  • feruloyl esterase
  • Glycoside Hydrolases
  • alpha-N-arabinofuranosidase