Simulations of cellulose translocation in the bacterial cellulose synthase suggest a regulatory mechanism for the dimeric structure of cellulose.
Simulations of cellulose translocation in the bacterial cellulose synthase suggest a regulatory mechanism for the dimeric structure of cellulose.
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DOI：
10.1039/c5sc04558d
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发表时间：
2016-05-01
期刊：
影响因子：
8.4
通讯作者：
Beckham GT
中科院分区：
文献类型：
--
作者：
Knott BC;Crowley MF;Himmel ME;Zimmer J;Beckham GT
除了提出一种调节纤维素结构的机制外，分子模拟还表明易位对纤维素生物合成不是限速的。细菌纤维素合酶（Bcs）的进行性循环包括将单个葡萄糖部分添加到生长的纤维素链的末端，随后新生链穿过质膜移位。这种易位的机制及其在进行性循环中的精确位置还不清楚。特别是，其基本结构单元是二聚体（纤维二糖）的聚合物（纤维素）如何通过一次添加一个单体（葡萄糖）来构建的分子细节尚未阐明。在这里，我们利用分子动力学模拟和自由能计算来阐明这些问题。我们发现，由一个葡萄糖单元的易位向前是非常有利的积极，给大于10千卡mol-1的自由能稳定。此外，只有一个小的障碍，易位，这意味着易位是不是速率限制内的Bcs进行周期（给定的实验速率纤维素合成在体外）。也许最重要的是，我们的研究结果还表明，在跨膜隧道入口的空间限制调节纤维素的二聚体结构。也就是说，当葡萄糖分子以与受体葡萄糖相同的取向添加到纤维素链时，末端葡萄糖在向前运动时自由旋转，从而表明纤维素的二聚体结构的调节机制。我们表征了保守和非保守的酶-多糖相互作用，驱动易位，并发现20的25个残基，强烈相互作用的易位纤维素链的模拟是保守的，主要是极性或芳香族侧链。我们的研究结果还允许一个动态分析的作用，所谓的“手指螺旋”纤维素易位，已观察到的结构。两者合计，这些研究结果有助于阐明的BCS进行性循环的易位步骤，并可能是广泛相关的多糖合成或降解酶，耦合催化与链易位。
In addition to suggesting a mechanism for regulating cellulose structure, molecular simulations indicate translocation is not rate-limiting for cellulose biosynthesis. The processive cycle of the bacterial cellulose synthase (Bcs) includes the addition of a single glucose moiety to the end of a growing cellulose chain followed by the translocation of the nascent chain across the plasma membrane. The mechanism of this translocation and its precise location within the processive cycle are not well understood. In particular, the molecular details of how a polymer (cellulose) whose basic structural unit is a dimer (cellobiose) can be constructed by adding one monomer (glucose) at a time are yet to be elucidated. Here, we have utilized molecular dynamics simulations and free energy calculations to the shed light on these questions. We find that translocation forward by one glucose unit is quite favorable energetically, giving a free energy stabilization of greater than 10 kcal mol–1. In addition, there is only a small barrier to translocation, implying that translocation is not rate limiting within the Bcs processive cycle (given experimental rates for cellulose synthesis in vitro). Perhaps most significantly, our results also indicate that steric constraints at the transmembrane tunnel entrance regulate the dimeric structure of cellulose. Namely, when a glucose molecule is added to the cellulose chain in the same orientation as the acceptor glucose, the terminal glucose freely rotates upon forward motion, thus suggesting a regulatory mechanism for the dimeric structure of cellulose. We characterize both the conserved and non-conserved enzyme–polysaccharide interactions that drive translocation, and find that 20 of the 25 residues that strongly interact with the translocating cellulose chain in the simulations are well conserved, mostly with polar or aromatic side chains. Our results also allow for a dynamical analysis of the role of the so-called ‘finger helix’ in cellulose translocation that has been observed structurally. Taken together, these findings aid in the elucidation of the translocation steps of the Bcs processive cycle and may be widely relevant to polysaccharide synthesizing or degrading enzymes that couple catalysis with chain translocation.