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The Pseudomonas cellulosa glycoside hydrolase family 51 arabinofuranosidase exhibits wide substrate specificity

Lookup NU author(s): Marie-helene Beylot, Dr Vincent McKie, Professor Harry Gilbert

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Abstract

To investigate the mechanism by which Pseudomonas cellulosa releases arabinose from polysaccharides and oligosaccharides, a gene library of P. cellulosa genomic DNA was screened for 4-methylumbelliferyl-α-L-arabinofuranosidase (MUAase) activity. A single MUAase gene (abf51A) was isolated, which encoded a non-modular glycoside hydrolase family (GH) 51 arabinofuranosidase (Abf51A) of 57000 Da. The substrate specificity of the Abf51A showed that it preferentially removed α1,2- and α1,3-linked arabinofuranose side chains from either arabinan or arabinoxylan, and hydrolysed α1,5-linked arabino-oligosaccharides, although at a much lower rate. The activity of Abf51A against arabinoxylan was similar to a GH62 arabinofuranosidase encoded by a P. cellulosa gene. Glu-194 and Glu321 of Abf51A are conserved in GH51 enzymes, and it has been suggested that these amino acids comprise the key catalytic acid/base and nucleophile residues, respectively. To evaluate this hypothesis the biochemical properties of E194A and E321A mutants of Abf51A were evaluated. The data were consistent with the view that Glu-194 and Glu-321 comprise the key catalytic residues of Abf51A. These data, in conjunction with the results presented in the accompanying paper [Beylot, Emami, McKie, Gilbert and Pell (2001) Biochem. J. 358, 599-605], indicate that P. cellulosa expresses a membrane-bound GH51 arabinofuranosidase that plays a pivotal role in releasing arabinose from a range of polysaccharides and oligosaccharides.


Publication metadata

Author(s): Beylot M-H, McKie VA, Voragen AGJ, Doeswijk-Voragen CHL, Gilbert HJ

Publication type: Article

Publication status: Published

Journal: Biochemical Journal

Year: 2001

Volume: 358

Issue: 3

Pages: 607-614

ISSN (print): 0264-6021

ISSN (electronic): 1470-8728

Publisher: Portland Press Ltd

URL: http://dx.doi.org/10.1042/0264-6021:3580607

DOI: 10.1042/0264-6021:3580607

PubMed id: 11535122


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