Molecular modeling of averaged rigor crossbridges from tomograms of insect flight muscle
Molecular modeling of averaged rigor crossbridges from tomograms of insect flight muscle
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DOI：
10.1016/s1047-8477(02)00013-8
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发表时间：
2002-04-01
影响因子：
3
通讯作者：
Taylor, KA
中科院分区：
文献类型：
--
作者：
Chen, LF;Winkler, H;Taylor, KA
电子断层扫描、对应分析、分子模型构建和实空间精修提供了无核苷酸状态（僵直态）下原位肌球蛋白横桥的详细三维结构，这种状态被认为代表了动力冲程的结束。来自昆虫飞行肌25纳米纵向切片的未平均断层图像保留了天然的结构变异。沿肌动蛋白丝每38.7纳米重复出现的横桥基序从断层图像中提取出来，并通过对应分析分类为25个类别平均值，这提高了信噪比。基于肌动蛋白和肌球蛋白亚片段1的原子结构的模型被重新构建以拟合11个类别平均值。应用实空间精修程序对重建进行定量拟合，并使拟合过程中引入的结构域之间的空间冲突最小化。这些综合程序表明，没有单一的肌球蛋白头部结构能够拟合所有的原位横桥。这种方法的有效性得到了以下支持：这些原子模型与来自脊椎动物肌肉的荧光探针数据以及与平滑肌重酶解肌球蛋白头部结合肌动蛋白时调节轻链交联的数据一致。（C）2002年爱思唯尔科学（美国）。保留所有权利。
Electron tomography, correspondence analysis, molecular model building, and real-space refinement provide detailed 3-D structures for in situ myosin crossbridges in the nucleotide-free state (rigor), thought to represent the end of the power stroke. Unaveraged tomograms from a 25-nm longitudinal section of insect flight muscle preserved native structural variation. Recurring crossbridge motifs that repeat every 38.7 nm along the actin filament were extracted from the tomogram and classified by correspondence analysis into 25 class averages, which improved the signal to noise ratio. Models based on the atomic structures of actin and of myosin subfragment 1 were rebuilt to fit 11 class averages. A real-space refinement procedure was applied to quantitatively fit the reconstructions and to minimize steric clashes between domains introduced during the fitting. These combined procedures show that no single myosin head structure can fit all the in situ crossbridges. The validity of the approach is supported by agreement of these atomic models with fluorescent probe data from vertebrate muscle as well as with data from regulatory light chain crosslinking between heads of smooth muscle heavy meromyosin when bound to actin. (C) 2002 Elsevier Science (USA). All rights reserved.