Department of Chemistry, The University of Chicago, Chicago, Illinois 60637, United States.
James Franck Institute, The University of Chicago, Chicago, Illinois 60637, United States.
Biochemistry. 2021 Oct 26;60(42):3125-3136. doi: 10.1021/acs.biochem.1c00583. Epub 2021 Oct 12.
Experimental evidence suggests that monomeric insulin exhibits significant conformational heterogeneity, and modifications of apparently disordered regions affect both biological activity and the longevity of pharmaceutical formulations, presumably through receptor binding and fibrillation/degradation, respectively. However, a microscopic understanding of conformational heterogeneity has been lacking. Here, we integrate all-atom molecular dynamics simulations with an analysis pipeline to investigate the structural ensemble of human insulin monomers. We find that 60% of the structures present at least one of the following elements of disorder: melting of the A-chain N-terminal helix, detachment of the B-chain N-terminus, and detachment of the B-chain C-terminus. We also observe partial melting and extension of the B-chain helix and significant conformational heterogeneity in the region containing the B-chain β-turn. We then estimate hydrogen-exchange protection factors for the sampled ensemble and find them in line with experimental results for KP-insulin, although the simulations underestimate the importance of unfolded states. Our results help explain the ready exchange of specific amide sites that appear to be protected in crystal structures. Finally, we discuss the implications for insulin function and stability.
实验证据表明，单体胰岛素表现出显著的构象异质性，而明显无序区域的修饰会影响生物活性和药物制剂的寿命，这可能分别通过受体结合和纤维化/降解来实现。然而，对于构象异质性的微观理解一直存在不足。在这里，我们整合了全原子分子动力学模拟和分析管道，以研究人胰岛素单体的结构整体。我们发现，60%的结构至少存在以下无序元素之一：A 链 N 端螺旋的熔化、B 链 N 端的脱离以及 B 链 C 端的脱离。我们还观察到 B 链螺旋的部分熔化和延伸，以及包含 B 链β转角的区域的显著构象异质性。然后，我们对采样整体的氢交换保护因子进行了估计，并发现它们与 KP 胰岛素的实验结果一致，尽管模拟低估了无规状态的重要性。我们的结果有助于解释在晶体结构中似乎受到保护的特定酰胺位点的快速交换。最后，我们讨论了对胰岛素功能和稳定性的影响。
