Cello-oligomer-binding dynamics and directionality in family 4 carbohydrate-binding modules
Cello-oligomer-binding dynamics and directionality in family 4 carbohydrate-binding modules
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DOI：
10.1093/glycob/cwv048
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发表时间：
2015-10-01
期刊：
影响因子：
4.3
通讯作者：
Payne, Christina M.
中科院分区：
文献类型：
--
作者：
Kognole, Abhishek A.;Payne, Christina M.
碳水化合物结合模块(CBMs)在调节纤维素酶的功能和了解CBMs选择性结合底物的蛋白质-碳水化合物识别机制方面发挥着重要作用，这对开发增强型生物质转化技术至关重要。CBM表现出有限的特异性，并且似乎以一种定向的方式结合多糖，这种结合方式取决于环氧相对于蛋白质折叠的位置。Cellulomonas FIMI Cel9B的两个家族4 CBM(CfCBM4)被报道优先结合纤维底物。然而，实验证据表明，这些CBM可能不会表现出对特定取向的热力学偏好。我们使用分子动力学(MD)和自由能计算来研究CfCBM4-1和CfCBM4-2中蛋白质-碳水化合物的识别机制，并阐明优先的配体结合方向。我们评估了四种纤维五糖的取向，包括晶体结构的取向和核磁共振(NMR)提出的其他三种取向。这四种取向根据配体还原端(RE)和吡喃糖环相对于蛋白质核心的位置而不同。分子动力学模拟表明，可能的取向减少为两种构象。计算的配体结合自由能识别出每一种取向都是同样有利的。计算的自由能与文献中的等温滴定量热法测量结果符合得很好。MD模拟进一步揭示了低聚糖相对于氨基酸的近似结构对称性，这在配体结合的混杂中起到了作用。一项对配体结合结构的调查表明，这种现象可能是属于β-三明治折叠的更广泛类别的蛋白质的特征。
Carbohydrate-binding modules (CBMs) play significant roles in modulating the function of cellulases, and understanding the protein-carbohydrate recognition mechanisms by which CBMs selectively bind substrate is critical to development of enhanced biomass conversion technology. CBMs exhibit a limited range of specificity and appear to bind polysaccharides in a directional fashion dictated by the position of the ring oxygen relative to the protein fold. The two family 4 CBMs of Cellulomonas fimi Cel9B (CfCBM4) are reported to preferentially bind cellulosic substrates. However, experimental evidence suggests that these CBMs may not exhibit a thermodynamic preference for a particular orientation. We use molecular dynamics (MD) and free energy calculations to investigate protein-carbohydrate recognition mechanisms in CfCBM4-1 and CfCBM4-2 and to elucidate preferential ligand-binding orientation. We evaluate four cellopentaose orientations including that of the crystal structure and three others suggested by nuclear magnetic resonance (NMR). These four orientations differ based on position of the ligand reducing end (RE) and pyranose ring orientations relative to the protein core. MD simulations indicate that the plausible orientations reduce to two conformations. Calculated ligand-binding free energy discerns each of the orientations is equally favorable. The calculated free energies are in excellent agreement with isothermal titration calorimetry measurements from the literature. MD simulations further reveal the approximate structural symmetry of the oligosaccharides relative to the amino acids along the binding cleft plays a role in the promiscuity of ligand binding. A survey of ligand-bound structures suggests this phenomenon may be characteristic of the broader class of proteins belonging to the beta-sandwich fold.