Department of Chemical and Biological Engineering, Drexel University, Philadelphia, Pennsylvania 19104, USA.
Proteins. 2010 May 1;78(6):1531-43. doi: 10.1002/prot.22670.
The insulin receptor (IR) is a homo-dimeric, disulfide-linked, membrane-spanning tyrosine kinase. IR displays negative cooperativity in insulin binding to its two pockets, suggesting "see-sawing" between symmetry-inverted conformations. The crystal structure of the dimeric IR ectodomain, IRDeltabeta [PDB code 2DTG (McKern et al., Nature 2006 443:218-221)], provides structural bases for this speculation. Unfortunately, neither binding pocket of the crystallographic IRDeltabeta structure allows steric accommodation of insulin. During almost 70-ns of all-atom, explicit-water MD simulation ( approximately 0.5 million atoms), IRDeltabeta undergoes significant asymmetric interdomain and intersubunit conformational fluctuations that do not alter its quaternary structure. Subtle variations in intersubunit buried surface area coincide with these conformational fluctuations, resulting in one easily-accessible insulin binding pocket with the other blocked. We use a combination of Metropolis Monte-Carlo and MD simulations to dock both T- and R-state insulin into the open binding pocket. Both complexes remain stable during 30-ns of MD simulation. In these complexes, "hexamer interface" residues on insulin directly contact the "site-2" epitope on the first type-III fibronectin domain (F1) of IR. Our results support the hypothesis that intersubunit flexibility of IR, governed by alternating modulation of buried intersubunit surface area, is the physical mechanism underlying a "see-saw" model of negative cooperativity.
胰岛素受体（IR）是一种同二聚体、二硫键连接的跨膜酪氨酸激酶。IR 在胰岛素与其两个口袋的结合中显示出负协同性，表明在对称反转构象之间存在“跷跷板”运动。二聚体 IR 胞外结构域（IRDeltabeta）的晶体结构[PDB 代码 2DTG（McKern 等人，自然 2006 年 443:218-221）]，为这种推测提供了结构基础。不幸的是，晶体结构的 IRDeltabeta 中没有一个结合口袋可以容纳胰岛素。在近 70-ns 的全原子、显式水 MD 模拟（约 50 万个原子）中，IRDeltabeta 经历了显著的不对称的域间和亚基间构象波动，而不会改变其四聚体结构。亚基间埋藏表面积的细微变化与这些构象波动一致，导致一个易于接近的胰岛素结合口袋，而另一个被阻断。我们使用 Metropolis 蒙特卡罗和 MD 模拟的组合将 T-和 R-状态胰岛素对接入开放的结合口袋。在 30-ns 的 MD 模拟中，两个复合物都保持稳定。在这些复合物中，胰岛素上的“六联体界面”残基直接与 IR 的第一个 III 型纤维连接蛋白结构域（F1）上的“位点 2”表位接触。我们的结果支持这样的假设，即 IR 的亚基间灵活性由埋藏亚基间表面积的交替调制控制，是负协同性“跷跷板”模型的物理机制。
