In order to elucidate how phosphatidylserine (PS6) interacts with AQP5 in a cell membrane, we developed a hybrid steered molecular dynamics (hSMD) method that involved: (1) Simultaneously steering two centers of mass of two selected segments of the ligand, and (2) equilibrating the ligand-protein complex with and without biasing the system. Validating hSMD, we first studied vascular endothelial growth factor receptor 1 (VEGFR1) in complex with N-(4-Chlorophenyl)-2-((pyridin-4-ylmethyl)amino)benzamide (8ST), for which the binding energy is known from in vitro experiments. In this study, our computed binding energy well agreed with the experimental value. Knowing the accuracy of this hSMD method, we applied it to the AQP5-lipid-bilayer system to answer an outstanding question relevant to AQP5’s physiological function: Will the PS6, a lipid having a single long hydrocarbon tail that was found in the central pore of the AQP5 tetramer crystal, actually bind to and inhibit AQP5’s central pore under near-physiological conditions, namely, when AQP5 tetramer is embedded in a lipid bilayer? We found, in silico, using the CHARMM 36 force field, that binding PS6 to AQP5 was a factor of 3 million weaker than “binding” it in the lipid bilayer. This suggests that AQP5’s central pore will not be inhibited by PS6 or a similar lipid in a physiological environment.

为了阐明磷脂酰丝氨酸（PS6）如何与细胞膜中的AQP5相互作用，我们开发了一种混合操纵分子动力学（hSMD）方法，该方法涉及：（1）同时操纵配体两个选定链段的两个质心，并且（ 2）在有和没有系统偏差的情况下平衡配体-蛋白质复合物。为了验证hSMD，我们首先研究了与N-（4-氯苯基）-2-（（吡啶-4-基甲基）氨基）苯甲酰胺（8ST）形成复合物的血管内皮生长因子受体1（VEGFR1），其结合能已知从体外实验。在这项研究中，我们计算的结合能与实验值非常吻合。知道此hSMD方法的准确性后，我们将其应用于AQP5-脂质-双层系统，以回答与AQP5的生理功能有关的一个悬而未决的问题：PS6，一种具有单个长烃尾的脂质会在AQP5的中央孔中被发现吗？ AQP5四聚体晶体在接近生理条件下，即当AQP5四聚体包埋在脂质双层中时，实际上结合并抑制AQP5的中央孔吗？我们在计算机上使用CHARMM 36力场在计算机上发现，将PS6与AQP5的结合比将其与脂质双层中的“结合”弱300万。这表明在生理环境中，PS6或类似脂质不会抑制AQP5的中央孔。