Improving the Estimation of pK_a Values for All Titratable Amino Acids at the Water/Membrane Interface

Understanding how pH-sensitive molecules behave when incorporated into lipid bilayers is a challenging problem that requires specialized methods and robust sampling. Techniques based on constant-pH molecular dynamics (CpHMD) can be employed to investigate the effects of pH on molecules at the water-membrane interface. Here, we compare the performance of enhanced sampling protocols coupled with CpHMD in describing protonation and pKa values of model Ala pentapeptide systems inserting into a DMPC membrane. From regular CpHMD, replica exchange (pHRE), and umbrella sampling (US-CpHMD), the latter provides more reliable and complete pKa profiles along bilayer insertion, in particular, at deeper regions of the membrane. Overall, there is a consistent trend for pKa values to shift toward the molecules’ neutral state, a crucial effect that enables a (de)protonation-assisted increase in membrane permeability. Building on US-CpHMD, we have successfully coupled an umbrella replica exchange scheme (REUS), which significantly improved the reliability of results for our Asp pentapeptide by enhancing configurational sampling between the umbrellas, albeit at a substantial computational cost.解析pH敏感分子嵌入脂质双分子层时的行为模式是一项极具挑战性的课题，亟需专用研究方法与可靠的采样策略。基于恒pH分子动力学（constant-pH molecular dynamics, CpHMD）的技术可用于探究pH对水-膜界面处分子的影响。本研究对比了耦合恒pH分子动力学的各类增强采样方案，在描述嵌入二肉豆蔻酰磷脂酰胆碱（DMPC）膜的丙氨酸（Ala）模型五肽体系的质子化状态与pKa值时的表现。结果表明，相较于常规恒pH分子动力学、pH副本交换（pHRE）方法，伞形采样-恒pH分子动力学（US-CpHMD）能够提供更可靠且完整的跨双分子层插入过程的pKa分布曲线，尤其在膜的深层区域表现更优。整体而言，pKa值呈现向分子中性态偏移的一致趋势，这一关键效应可实现脱/质子化辅助的膜通透性提升。基于伞形采样-恒pH分子动力学框架，本研究进一步成功耦合了伞形副本交换方案（REUS），该方案通过增强伞形窗口间的构象采样，显著提升了天冬氨酸（Asp）五肽体系研究结果的可靠性，尽管其计算成本较高。