Structural and biochemical characterization of the broad substrate specificity of Bacteroides thetaiotaomicron commensal sialidase.

Kwang-Hyun Park, Min-Gyu Kim, Hee-Jeong Ahn, Dae-Han Lee, Jin-Hyo Kim, Young-Wan Kim, Eui-Jeon Woo

Index: Biochim. Biophys. Acta 1834(8) , 1510-9, (2013)

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Abstract

Sialidases release the terminal sialic acid residue from a wide range of sialic acid-containing polysaccharides. Bacteroides thetaiotaomicron, a symbiotic commensal microbe, resides in and dominates the human intestinal tract. We characterized the recombinant sialidase from B. thetaiotaomicron (BTSA) and demonstrated that it has broad substrate specificity with a relative activity of 97, 100 and 64 for 2,3-, 2,6- and 2,8-linked sialic substrates, respectively. The hydrolysis activity of BTSA was inhibited by a transition state analogue, 2-deoxy-2,3-dehydro-N-acetyl neuraminic acid, by competitive inhibition with a Ki value of 35μM. The structure of BSTA was determined at a resolution of 2.3Å. This structure exhibited a unique carbohydrate-binding domain (CBM) at its N-terminus (a.a. 23-190) that is adjacent to the catalytic domain (a.a. 191-535). The catalytic domain has a conserved arginine triad with a wide-open entrance for the substrate that exposes the catalytic residue to the surface. Unlike other pathogenic sialidases, the polysaccharide-binding site in the CBM is near the active site and possibly holds and positions the polysaccharide substrate directly at the active site. The structural feature of a wide substrate-binding groove and closer proximity of the polysaccharide-binding site to the active site could be a unique signature of the commensal sialidase BTSA and provide a molecular basis for its pharmaceutical application. Copyright © 2013 Elsevier B.V. All rights reserved.