The biochemical kinetics underlying actin movement generated by one and many skeletal muscle myosin molecules
The biochemical kinetics underlying actin movement generated by one and many skeletal muscle myosin molecules
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DOI：
10.1016/s0006-3495(02)75560-4
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发表时间：
2002-04-01
影响因子：
3.4
通讯作者：
Warshaw, DM
中科院分区：
文献类型：
--
作者：
Baker, JE;Brosseau, C;Warshaw, DM
为了更好地理解骨骼肌肌球蛋白分子如何移动肌动蛋白丝，我们确定了单个肌球蛋白分子产生运动的生物化学过程，并研究它如何与一群肌球蛋白分子产生运动的生物化学过程相关联。首先，通过测量各种配体（ATP、ADP和Pi）对激光阱中事件持续时间τ（on）的影响，我们确定了单个肌球蛋白分子产生的肌动蛋白丝逐步移动所依据的生物化学动力学。接下来，通过测量这些相同配体对体外运动测定中肌动蛋白速度V的影响，我们确定了一群肌球蛋白分子产生的肌动蛋白丝连续移动所依据的生物化学过程。观察到的Pi对单分子机械化学的影响表明，单个肌球蛋白分子产生运动与肌动蛋白诱导的Pi解离密切相关。我们通过测量由平滑肌HMM突变（导致β释放速率降低）引起的单分子机械化学变化，为这种关系获得了额外的证据。相比之下，我们观察到一群肌球蛋白分子产生运动在低[ATP]时受ATP诱导的肌动蛋白解离限制（即V随1/τ（on）变化），但在高[ATP]时偏离这种关系。单分子数据独特地提供了在最小负载下肌动球蛋白ATP酶反应的基本机械化学的直接测量，并作为一个模型的明确基础，在该模型中，与肌动蛋白相连的肌球蛋白分子施加负载。
To better understand how skeletal muscle myosin molecules move actin filaments, we determine the motion-generating biochemistry of a single myosin molecule and study how it scales with the motion-generating biochemistry of an ensemble of myosin molecules. First, by measuring the effects of various ligands (ATP, ADP, and P-i) on event lifetimes, tau(on), in a laser trap, we determine the biochemical kinetics underlying the stepwise movement of an actin filament generated by a single myosin molecule. Next, by measuring the effects of these same ligands on actin velocities, V, in an in vitro motility assay, we determine the biochemistry underlying the continuous movement of an actin filament generated by an ensemble of myosin molecules. The observed effects of P-i on single molecule mechanochemistry indicate that motion generation by a single myosin molecule is closely associated with actin-induced P-i dissociation. We obtain additional evidence for this relationship by measuring changes in single molecule mechanochemistry caused by a smooth muscle HMM mutation that results in a reduced beta-release rate. In contrast, we observe that motion generation by an ensemble of myosin molecules is limited by ATIP-induced actin dissociation (i.e., V varies as 1/tau(on)) at low [ATP], but deviates from this relationship at high [ATP]. The single-molecule data uniquely provide a direct measure of the fundamental mechanochemistry of the actomyosin ATPase reaction under a minimal load and serve as a clear basis for a model of ensemble motility in which actin-attached myosin molecules impose a load.