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  • Critical roles of Asp270 and Trp273 in the α-repeat of the carbohydrate-binding module of endo-1,3-β-glucanase for laminarin-binding avidity.

Critical roles of Asp270 and Trp273 in the α-repeat of the carbohydrate-binding module of endo-1,3-β-glucanase for laminarin-binding avidity.

Glycoconjugate journal (2011-12-27)
Tomonari Tamashiro, Yoichi Tanabe, Teikichi Ikura, Nobutoshi Ito, Masayuki Oda
ABSTRACT

A carbohydrate-binding module from family 13 (CBM13), appended to the catalytic domain of endo-1,3-β-glucanase from Cellulosimicrobium cellulans, was overexpressed in E. coli, and its interactions with β-glucans, laminarin and laminarioligosaccharides, were analyzed using surface plasmon resonance biosensor and isothermal titration calorimetry. The association constants for laminarin and laminarioligosaccharides were determined to be approximately 10(6) M(-1) and 10(4) M(-1), respectively, indicating that 2 or 3 binding sites in the α-, β-, and γ-repeats of CBM13 are involved in laminarin binding in a cooperative manner. The binding avidity is approximately 2-orders higher than the monovalent binding affinity. Mutational analysis of the conserved Asp residues in the respective repeats showed that the α-repeat primarily contributes to β-glucan binding. A Trp residue is predicted to be exposed to the solvent only in the α-repeat and would contribute to β-glucan binding. The α-repeat bound β-glucan with an affinity of approximately 10(4) M(-1), and the other repeats additionally bound laminarin, resulting in the increased binding avidity. This binding is unique compared to the recognition mode of another CBM13 from Streptomyces lividans xylanase.

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Sigma-Aldrich
β-(1→3)-D-Glucanase from Helix pomatia, ≥0.2 U/mg
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