Characterization of two distinct feruloyl esterases, AoFaeB and AoFaeC, from Aspergillus oryzae

Appl Microbiol Biotechnol. 2009 Jun;83(4):689-96. doi: 10.1007/s00253-009-1913-z. Epub 2009 Feb 26.

Abstract

Two hypothetical proteins XP_001818628 and XP_001819091 (designated AoFaeB and AoFaeC, respectively), showing sequence identity with known type-C feruloyl esterases, have been found in the genomic sequence of Aspergillus oryzae. We cloned the putative A. oryzae feruloyl esterase-encoding genes and expressed them in Pichia pastoris. Both purified recombinant AoFaeB (rAoFaeB) and AoFaeC (rAoFaeC) had apparent relative molecular masses of 61,000 and 75,000, respectively, on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. After N-deglycosylation, both proteins had a relative molecular mass of 55,000. The optimum pH for rAoFaeB was 6.0, although it was stable at pH values ranging from 3.0 to 9.0; rAoFaeC had an optimum pH of 6.0 and was stable in the pH range of 7.0-10.0. Thermostability of rAoFaeC was greater than that of rAoFaeB. Whereas rAoFaeC displayed hydrolytic activity toward methyl caffeate, methyl p-coumarate, methyl ferulate, and methyl sinapate, rAoFaeB displayed hydrolytic activity toward methyl caffeate, methyl p-coumarate, and methyl ferulate but not toward methyl sinapate. Substrate specificity profiling of rAoFaeB and rAoFaeC revealed type-B and type-C feruloyl esterases, respectively. Ferulic acid was efficiently released from wheat arabinoxylan when both esterases were applied with xylanase from Thermomyces lanuginosus. Both recombinant proteins also exhibited hydrolytic activity toward chlorogenic acid.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Aspergillus oryzae / enzymology*
  • Aspergillus oryzae / genetics
  • Aspergillus oryzae / metabolism
  • Carboxylic Ester Hydrolases / chemistry
  • Carboxylic Ester Hydrolases / genetics
  • Carboxylic Ester Hydrolases / isolation & purification*
  • Carboxylic Ester Hydrolases / metabolism*
  • Chlorogenic Acid / metabolism
  • Cloning, Molecular
  • Coumaric Acids / metabolism
  • Enzyme Stability
  • Fungal Proteins / chemistry
  • Fungal Proteins / genetics
  • Fungal Proteins / isolation & purification*
  • Fungal Proteins / metabolism*
  • Gene Expression
  • Hydrogen-Ion Concentration
  • Molecular Sequence Data
  • Molecular Weight
  • Pichia / genetics
  • Recombinant Proteins / genetics
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Sequence Alignment
  • Substrate Specificity
  • Temperature
  • Xylans / metabolism

Substances

  • Coumaric Acids
  • Fungal Proteins
  • Recombinant Proteins
  • Xylans
  • Chlorogenic Acid
  • arabinoxylan
  • ferulic acid
  • Carboxylic Ester Hydrolases
  • feruloyl esterase