| Summary: | 碩士 === 國立臺北科技大學 === 有機高分子研究所 === 98 === Fibrobacter succinogenes 1,3-1,4-β-D-glucanase hydrolyzes and cleaves β-1,4-glycosidic bonds precisely where β-1,3-glycosidic linkages are located prior to β-1,4-glycosidic linkages in lichenan or β-D-glucans. Four aromatic residues, Phe40, Tyr42, Trp203 and Phe205 in the catalytic domain of Fibrobacter succinogenes 1,3-1,4-β-D-glucanase (TFs-β-glucanase) interact with sugar units of the product via hydrophobic stacking interactions. For the purpose of understanding of the structural and functional roles of the active site residues, many mutants have been characterized. The crystal structures of the F40I and W203F mutants revealed that two extra calcium ions and a Tris molecule have been identified. The Tris molecule was found at the position normally taken by substrate binding to the -1 subsite and bound to the catalytic residues Glu56 and Glu60. In addition, a second Ca2+ ion was found near residues Phe152 and Glu154, and a third near the active site. In order to understand the kinetic inhibition properties of Tris and the calcium ion, kinetic experiments were performed. Further kinetics analysis revealed that Tris is a competitive inhibitor of the enzyme, while calcium ion is a non-competitive inhibitor.
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