Singh Abhishek, Hitchcock-DeGregori Sarah E
Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
Structure. 2006 Jan;14(1):43-50. doi: 10.1016/j.str.2005.09.016.
The coiled coil is a widespread motif involved in oligomerization and protein-protein interactions, but the structural requirements for binding to target proteins are poorly understood. To address this question, we measured binding of tropomyosin, the prototype coiled coil, to actin as a model system. Tropomyosin binds to the actin filament and cooperatively regulates its function. Our results support the hypothesis that coiled-coil domains that bind to other proteins are flexible. We made mutations that alter interface packing and stability as well as mutations in surface residues in a postulated actin binding site. Actin affinity, measured by cosedimentation, was correlated with coiled-coil stability and local instability and side chain flexibility, analyzed with circular dichroism and fluorescence spectroscopy. The flexibility from interruptions in the stable coiled-coil interface is essential for actin binding. The surface residues in a postulated actin binding site participate in actin binding when the coiled coil within it is poorly packed.
卷曲螺旋是一种广泛存在的基序，参与寡聚化和蛋白质-蛋白质相互作用，但对其与靶蛋白结合的结构要求却知之甚少。为了解决这个问题，我们以原肌球蛋白（卷曲螺旋的原型）与肌动蛋白的结合作为模型系统进行了测量。原肌球蛋白与肌动蛋白丝结合并协同调节其功能。我们的结果支持这样一种假设，即与其他蛋白质结合的卷曲螺旋结构域是灵活的。我们进行了改变界面堆积和稳定性的突变，以及在假定的肌动蛋白结合位点的表面残基处进行突变。通过共沉降测量的肌动蛋白亲和力与卷曲螺旋稳定性、局部不稳定性以及用圆二色光谱和荧光光谱分析的侧链灵活性相关。稳定的卷曲螺旋界面中断所带来的灵活性对于肌动蛋白结合至关重要。当假定的肌动蛋白结合位点内的卷曲螺旋堆积不佳时，该位点的表面残基会参与肌动蛋白结合。
