Family 39 alpha-l-iduronidases and beta-D-xylosidases react through similar glycosyl-enzyme intermediates: identification of the human iduronidase nucleophile

Biochemistry. 2003 Jul 8;42(26):8054-65. doi: 10.1021/bi034293v.


The inclusion of both beta-D-xylosidases and alpha-L-iduronidases within the same sequence-related family (family 39), despite the considerable difference in substrate structures and poor sequence conservation around the putative nucleophile, raises concerns about whether a common mechanism is followed by the two enzymes. A novel anchimeric assistance mechanism for iduronidases involving a lactone intermediate is one possibility. NMR analysis of the methanolysis reaction catalyzed by human alpha-L-iduronidase reveals that, as with the beta-D-xylosidases, alpha-L-iduronidase is a retaining glycosidase. Using two different mechanism-based inactivators, 5-fluoro-alpha-L-iduronyl fluoride and 2-deoxy-2-fluoro-alpha-L-iduronyl fluoride, the active site nucleophile in the human alpha-L-iduronidase was identified as Glu299 within the (295)IYNDEAD(301) sequence. The equivalent, though loosely predicted, glutamic acid was identified as the nucleophile in the family 39 beta-D-xylosidase from Bacillus sp. [Vocadlo, D., et al. (1998) Biochem. J. 335, 449-455]; thus, a common mechanism involving a covalent glycosyl-enzyme intermediate that adopts the rather uncommon (2,5)B conformation is predicted.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Bacillus / enzymology*
  • Binding Sites
  • Catalytic Domain
  • Conserved Sequence
  • Glutamic Acid / chemistry
  • Humans
  • Iduronic Acid / analogs & derivatives
  • Iduronic Acid / chemical synthesis
  • Iduronic Acid / metabolism
  • Iduronidase / chemistry*
  • Iduronidase / metabolism*
  • Kinetics
  • Mass Spectrometry / methods*
  • Molecular Sequence Data
  • Peptide Fragments / chemistry
  • Sequence Homology, Amino Acid
  • Stereoisomerism
  • Xylosidases / chemistry*
  • Xylosidases / metabolism*


  • Peptide Fragments
  • Iduronic Acid
  • Glutamic Acid
  • Xylosidases
  • exo-1,4-beta-D-xylosidase
  • Iduronidase