Flexibility within myosin heads revealed by negative stain and single-particle analysis.
Flexibility within myosin heads revealed by negative stain and single-particle analysis.
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DOI：
10.1083/jcb.139.3.675
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发表时间：
1997-11-03
期刊：
影响因子：
--
通讯作者：
Trinick J
中科院分区：
文献类型：
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作者：
Burgess SA;Walker ML;White HD;Trinick J
负染色肌球蛋白的电子显微镜先前已经揭示了分子头部内的三个离散区域。然而，尽管可能的分辨率为~ 2 nm，但很难在这些区域内直接识别一致的细节。这是由于头部构象和染色的变化。在这项研究中，我们采用单粒子图像处理，并将头部分类为均匀组。在对这些组进行平均后，改善的信噪比显示出大大改善的细节。将图像平均值与模拟亚片段-1 （S1）原子结构的阴性染色模型进行比较。这表明这三个头部区域对应于运动域和基本轻链和调节轻链。图像平均值与S1模型的特定视图非常相似。他们还揭示了运动和调节区域之间相当大的灵活性，尽管这些分子是在缺乏核苷酸的情况下制备的。这种灵活性可能是由于调节结构域围绕运动结构域的旋转，其中调节轻链的相对运动可达12纳米，并且在动画序列中最清楚地说明了这一点（可在http://www.leeds.ac.uk/chb/muscle/ myosinhead.html中获得）。S1原子模型的急剧弯曲构象在我们的数据中很少看到，更典型的是更直的头部。
Electron microscopy of negatively stained myosin has previously revealed three discrete regions within the heads of the molecule. However, despite a probable resolution of ∼2 nm, it is difficult to discern directly consistent details within these regions. This is due to variability in both head conformation and in staining. In this study, we applied single-particle image processing and classified heads into homogeneous groups. The improved signal-to-noise ratio after averaging these groups reveals substantially improved detail. The image averages were compared to a model simulating negative staining of the atomic structure of subfragment-1 (S1). This shows that the three head regions correspond to the motor domain and the essential and regulatory light chains. The image averages were very similar to particular views of the S1 model. They also revealed considerable flexibility between the motor and regulatory domains, despite the molecules having been prepared in the absence of nucleotide. This flexibility probably results from rotation of the regulatory domain about the motor domain, where the relative movement of the regulatory light chain is up to 12 nm, and is most clearly illustrated in animated sequences (available at http://www.leeds.ac.uk/chb/muscle/ myosinhead.html). The sharply curved conformation of the atomic model of S1 is seen only rarely in our data, with straighter heads being more typical.