Gorfe Alemayehu A, Chang Chia-en A, Ivanov Ivaylo, McCammon J Andrew
Department of Chemistry and Biochemistry, Center for Theoretical Biological Physics, University of California at San Diego, La Jolla, California 92093-0365, USA.
Biophys J. 2008 Feb 15;94(4):1144-54. doi: 10.1529/biophysj.107.117879. Epub 2007 Oct 5.
Acetylcholinesterase rapidly hydrolyzes the neurotransmitter acetylcholine in cholinergic synapses, including the neuromuscular junction. The tetramer is the most important functional form of the enzyme. Two low-resolution crystal structures have been solved. One is compact with two of its four peripheral anionic sites (PAS) sterically blocked by complementary subunits. The other is a loose tetramer with all four subunits accessible to solvent. These structures lacked the C-terminal amphipathic t-peptide (WAT domain) that interacts with the proline-rich attachment domain (PRAD). A complete tetramer model (AChEt) was built based on the structure of the PRAD/WAT complex and the compact tetramer. Normal mode analysis suggested that AChEt could exist in several conformations with subunits fluctuating relative to one another. Here, a multiscale simulation involving all-atom molecular dynamics and C alpha-based coarse-grained Brownian dynamics simulations was carried out to investigate the large-scale intersubunit dynamics in AChEt. We sampled the ns-mus timescale motions and found that the tetramer indeed constitutes a dynamic assembly of monomers. The intersubunit fluctuation is correlated with the occlusion of the PAS. Such motions of the subunits "gate" ligand-protein association. The gates are open more than 80% of the time on average, which suggests a small reduction in ligand-protein binding. Despite the limitations in the starting model and approximations inherent in coarse graining, these results are consistent with experiments which suggest that binding of a substrate to the PAS is only somewhat hindered by the association of the subunits.
乙酰胆碱酯酶可快速水解胆碱能突触（包括神经肌肉接头）中的神经递质乙酰胆碱。四聚体是该酶最重要的功能形式。已解析出两种低分辨率晶体结构。一种结构紧凑，其四个外周阴离子位点（PAS）中的两个在空间上被互补亚基阻断。另一种是松散的四聚体，所有四个亚基都可与溶剂接触。这些结构缺少与富含脯氨酸的附着结构域（PRAD）相互作用的C端两亲性t肽（WAT结构域）。基于PRAD/WAT复合物和紧凑四聚体的结构构建了一个完整的四聚体模型（AChEt）。正常模式分析表明，AChEt可以存在于几种构象中，亚基之间相对波动。在此，进行了一项涉及全原子分子动力学和基于Cα的粗粒化布朗动力学模拟的多尺度模拟，以研究AChEt中亚基间的大规模动力学。我们对纳秒至微秒时间尺度的运动进行了采样，发现四聚体确实构成了单体的动态组装体。亚基间的波动与PAS的封闭相关。亚基的这种运动“控制”配体与蛋白质的结合。这些“门”平均有超过80%的时间是打开的，这表明配体与蛋白质的结合略有减少。尽管起始模型存在局限性以及粗粒化中固有的近似性，但这些结果与实验一致，实验表明底物与PAS的结合仅在一定程度上受到亚基结合的阻碍。
