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How family 26 glycoside hydrolases orchestrate catalysis on different polysaccharides- Structure and activity of a Clostridium thermocellum lichenase, CtLic26A.

Taylor, E.J., Goyal, A., Guerreiro, C.I.P.D., Prates, J.A.M., Money, V.A., Ferry, N., Morland, C., Planas, A., Macdonald, J.A., Stick, R.V., Gilbert, H.J., Fontes, C.M.G.A. and Davies, G.J. (2005) How family 26 glycoside hydrolases orchestrate catalysis on different polysaccharides- Structure and activity of a Clostridium thermocellum lichenase, CtLic26A. Journal of Biological Chemistry, 280 (38). pp. 32761-32767. ISSN 0021-9258

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Abstract

One of the most intriguing features of the 90 glycoside hydrolase families (GHs) is the range of specificities displayed by different members of the same family, whereas the catalytic apparatus and mechanism are often invariant. Family GH26 predominantly comprises -1,4 mannanases; however, a bifunctional Clostridium thermocellum GH26 member (hereafter CtLic26A) displays a markedly different specificity. We show that CtLic26A is a lichenase, specific for mixed (Glc1,4Glc1,4Glc1,3)n oligo- and polysaccharides, and displays no activity on manno-configured substrates or -1,4-linked homopolymers of glucose or xylose. The three-dimensional structure of the native form of CtLic26A has been solved at 1.50-Å resolution, revealing a characteristic (/)8 barrel with Glu-109 and Glu-222 acting as the catalytic acid/base and nucleophile in a double-displacement mechanism. The complex with the competitive inhibitor, Glc--1,3-isofagomine (Ki 1 µM), at 1.60 Å sheds light on substrate recognition in the –2 and –1 subsites and illuminates why the enzyme is specific for lichenan-based substrates. Hydrolysis of -mannosides by GH26 members is thought to proceed through transition states in the B2,5 (boat) conformation in which structural distinction of glucosides versus mannosides reflects not the configuration at C2 but the recognition of the pseudoaxial O3 of the B2,5 conformation. We suggest a different conformational itinerary for the GH26 enzymes active on gluco-configured substrates.

Item Type: Article
Institution: The University of York
Academic Units: The University of York > Mathematics (York)
Depositing User: York RAE Import
Date Deposited: 23 Apr 2009 14:34
Last Modified: 23 Apr 2009 14:34
Published Version: http://dx.doi.org/10.1074/jbc.M506580200
Status: Published
Publisher: The American Society for Biochemistry and Molecular Biology
Identification Number: 10.1074/jbc.M506580200
URI: http://eprints.whiterose.ac.uk/id/eprint/6566

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