Analysis of keystone enzyme in Agar hydrolysis provides insight into the degradation (of a polysaccharide from) red seaweeds

J Biol Chem. 2012 Apr 20;287(17):13985-95. doi: 10.1074/jbc.M112.345645. Epub 2012 Mar 5.

Abstract

Agars are abundant polysaccharides from marine red algae, and their chemical structure consists of alternating D-galactose and 3,6-anhydro-L-galactose residues, the latter of which are presumed to make the polymer recalcitrant to degradation by most terrestrial bacteria. Here we study a family 117 glycoside hydrolase (BpGH117) encoded within a recently discovered locus from the human gut bacterium Bacteroides plebeius. Consistent with this locus being involved in agarocolloid degradation, we show that BpGH117 is an exo-acting 3,6-anhydro-α-(1,3)-L-galactosidase that removes the 3,6-anhydrogalactose from the non-reducing end of neoagaro-oligosaccharides. A Michaelis complex of BpGH117 with neoagarobiose reveals the distortion of the constrained 3,6-anhydro-L-galactose into a conformation that favors catalysis. Furthermore, this complex, supported by analysis of site-directed mutants, provides evidence for an organization of the active site and positioning of the catalytic residues that are consistent with an inverting mechanism of catalysis and suggests that a histidine residue acts as the general acid. This latter feature differs from the vast majority of glycoside hydrolases, which use a carboxylic acid, highlighting the alternative strategies that enzymes may utilize in catalyzing the cleavage of glycosidic bonds.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Agar / chemistry*
  • Binding Sites
  • Catalysis
  • Catalytic Domain
  • Chromatography, Thin Layer / methods
  • Cloning, Molecular
  • Crystallography, X-Ray / methods
  • Glycoside Hydrolases / chemistry*
  • Hydrolysis
  • Models, Molecular
  • Oligonucleotides / chemistry
  • Polysaccharides / chemistry
  • Protein Conformation
  • Protein Folding
  • Seaweed / chemistry*
  • Substrate Specificity

Substances

  • Oligonucleotides
  • Polysaccharides
  • Agar
  • Glycoside Hydrolases

Associated data

  • PDB/4AK5
  • PDB/4AK6
  • PDB/4AK7