Characterization of recombinant rhamnogalacturonan <i>α</i>-L-rhamnopyranosyl-(1,4)-<i>α</i>-D-galactopyranosyluronide lyase from <i>Aspergillus aculeatus</i>

  • Mutter M
  • Colquhoun I
  • Beldman G
 et al. 
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Abstract

The four major oligomeric reaction products from saponified modified hairy regions (MHR-S) from apple, produced by recombinant rhamnogalacturonan (RG) alpha-L-rhamnopyranosyl-(1, 4)-alpha-D-galactopyranosyluronide lyase (rRG-lyase) from Aspergillus aculeatus, were isolated and characterized by 1H-nuclear magnetic resonance spectroscopy. They contain an alternating RG backbone with a degree of polymerization of 4, 6, 8, and 10 and with an alpha-Delta-(4,5)-unsaturated D-galactopyranosyluronic acid at the nonreducing end and an L-rhamnopyranose at the reducing end. L-Rhamnopyranose units are substituted at C-4 with beta-galactose. The maximum reaction rate of rRG-lyase toward MHR-S at pH 6.0 and 31 degreesC was 28 units mg-1. rRG-lyase and RG-hydrolase cleave the same alternating RG I subunit in MHR. Both of these enzymes fragment MHR by a multiple attack mechanism. The catalytic efficiency of rRG-lyase for MHR increases with decreasing degree of acetylation. Removal of arabinose side chains improves the action of rRG-lyase toward MHR-S. In contrast, removal of galactose side chains decreased the catalytic efficiency of rRG-lyase. Native RG-lyase was purified from A. aculeatus, characterized, and found to be similar to the rRG-lyase expressed in Aspergillus oryzae.

Author-supplied keywords

  • Acetylation
  • Aspergillus
  • Aspergillus: chemistry
  • Aspergillus: enzymology
  • Carbohydrate Sequence
  • Enzyme Activation
  • Magnetic Resonance Spectroscopy
  • Molecular Conformation
  • Molecular Sequence Data
  • Pectins
  • Pectins: metabolism
  • Polysaccharide-Lyases
  • Polysaccharide-Lyases: chemistry
  • Polysaccharide-Lyases: genetics
  • Polysaccharide-Lyases: metabolism
  • Recombinant Proteins
  • Recombinant Proteins: chemistry
  • Recombinant Proteins: metabolism

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Authors

  • M Mutter

  • I J Colquhoun

  • G Beldman

  • H a Schols

  • E J Bakx

  • a G Voragen

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