Specificity and Affinity of Substrate Binding by a Family 17 Carbohydrate-Binding Module from Clostridium cellulovorans Cellulase 5A

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• 作者：Alisdair B. Boraston ; Patrick Chiu ; R. Antony J. Warren ; and Douglas G. Kilburn
• 刊名：Biochemistry
• 出版年：2000
• 出版时间：September 12, 2000
• 年：2000
• 卷：39
• 期：36
• 页码：11129 - 11136
• 全文大小：188K
• 年卷期：v.39,no.36(September 12, 2000)
• ISSN：1520-4995

文摘

The C-terminal carbohydrate-binding module (CBM17) from Clostridium cellulovorans cellulase5A is a -1,4-glucan binding module with a preference for soluble chains. CBM17 binds to phosphoricacid swollen Avicel (PASA) and Avicel with association constants of 2.9 (±0.2) × 10⁵ and 1.6 (±0.2)× 10⁵ M^-1, respectively. The capacity values for PASA and Avicel were 11.9 and 0.4 mol/g of cellulose,respectively. CBM17 did not bind to crystalline cellulose. CBM17 bound tightly to soluble barley -glucanand the derivatized celluloses HEC, EHEC, and CMC. The association constants for binding to barley-glucan, HEC, and EHEC were approximately 2.0 × 10⁵ M^-1. Significant binding affinities were foundfor cello-oligosaccharides greater than three glucose units in length. The affinities for cellotriose,cellotetraose, cellopentaose, and cellohexaose were 1.2 (±0.3) × 10³, 4.3 (±0.4) × 10³, 3.8 (±0.1) ×10⁴, and 1.5 (±0.0) × 10⁵ M^-1, respectively. Fluorescence quenching studies and N-bromosuccinimidemodification indicate the participation of tryptophan residues in ligand binding. The possible architectureof the ligand-binding site is discussed in terms of its binding specificity, affinity, and the participation oftryptophan residues.