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https://hdl.handle.net/2440/7022
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Type: | Journal article |
Title: | Family 39 a-L-iduronidases and b-D-xylosidases react through similar glycosyl-enzyme intermediates: Identification of the human iduronidase nucleophile |
Author: | Nieman, C. Wong, A. He, S. Clarke, L. Hopwood, J. Withers, S. |
Citation: | Biochemistry, 2003; 42(26):8054-8065 |
Publisher: | Amer Chemical Soc |
Issue Date: | 2003 |
ISSN: | 0006-2960 1520-4995 |
Abstract: | 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. |
Keywords: | Humans Bacillus Iduronic Acid Iduronidase Xylosidases Glutamic Acid Peptide Fragments Binding Sites Amino Acid Sequence Catalytic Domain Conserved Sequence Sequence Homology, Amino Acid Kinetics Stereoisomerism Molecular Sequence Data Mass Spectrometry |
DOI: | 10.1021/bi034293v |
Published version: | http://dx.doi.org/10.1021/bi034293v |
Appears in Collections: | Aurora harvest Paediatrics publications |
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