Department of Biochemistry and Biophysics, University of Rochester School of Medicine, Rochester, New York 14642, USA.
Proteins. 2011 Feb;79(2):518-27. doi: 10.1002/prot.22899.
Previous NMR experiments on unbound G protein βγ heterodimer suggested that particular residues in the binding interface are mobile on the nanosecond timescale. In this work we performed nanosecond-timescale molecular dynamics simulations to investigate conformational changes and dynamics of Gβγ in the presence of several binding partners: a high-affinity peptide (SIGK), phosducin, and the GDP-bound α subunit. In these simulations, the high mobility of GβW99 was reduced by SIGK, and it appeared that a tyrosine might stabilize GβW99 by hydrophobic or aromatic stacking interactions in addition to hydrogen bonds. Simulations of the phosducin-Gβγ complex showed that the mobility of GβW99 was restricted, consistent with inferences from NMR. However, large-scale conformational changes of Gβγ due to binding, which were hypothesized in the NMR study, were not observed in the simulations, most likely due to their short (nanosecond) duration. A pocket consisting of hydrophobic amino acids on Gα appears to restrict GβW99 mobility in the crystal structure of the Gαβγ? heterotrimer. The simulation trajectories are consistent with this idea. However, local conformational changes of residues GβW63, GβW211, GβW297, GβW332, and GβW339 were detected during the MD simulations. As expected, the magnitude of atomic fluctuations observed in simulations was greater for α than for the βγ subunits, suggesting that α has greater flexibility. These observations support the notion that to maintain the high mobility of GβW99 observed by solution NMR requires that the Gβ-α interface must open up on time scale longer than can be observed in nanosecond scale simulations.
先前针对非结合状态下 G 蛋白βγ异源二聚体的 NMR 实验表明，结合界面中的某些特定残基在纳秒时间尺度上是运动的。在这项工作中，我们进行了纳秒时间尺度的分子动力学模拟，以研究在几种结合配体存在的情况下 Gβγ的构象变化和动力学：一种高亲和力肽（SIGK）、磷蛋白和 GDP 结合的α亚基。在这些模拟中，SIGK 降低了 GβW99 的高迁移率，似乎一个酪氨酸可能通过疏水或芳香堆积相互作用以及氢键来稳定 GβW99。磷蛋白-Gβγ复合物的模拟表明，GβW99 的迁移率受到限制，这与 NMR 的推论一致。然而，由于结合而导致的 Gβγ的大规模构象变化，在 NMR 研究中被假设，但在模拟中没有观察到，最有可能是由于它们的持续时间短（纳秒）。在 Gαβγ 异三聚体的晶体结构中，一个由疏水性氨基酸组成的口袋似乎限制了 GβW99 的迁移率。模拟轨迹与这一观点一致。然而，在 MD 模拟过程中检测到残基 GβW63、GβW211、GβW297、GβW332 和 GβW339 的局部构象变化。正如预期的那样，在模拟中观察到的原子波动幅度对于α大于βγ 亚基，这表明α具有更大的灵活性。这些观察结果支持了这样一种观点，即要保持通过溶液 NMR 观察到的 GβW99 的高迁移率，需要 Gβ-α 界面在纳秒尺度模拟无法观察到的时间尺度上打开。
