Charged Residues Distribution Modulates Selectivity of the Open State of Human Isoforms of the Voltage Dependent Anion-Selective Channel

Voltage Dependent Anion-selective Channels (VDACs) are pore-forming proteins located in the outer mitochondrial membrane. They are responsible for the access of ions and energetic metabolites into the inner membrane transport systems. Three VDAC isoforms exist in mammalian, but their specific role is unknown. In this work we have performed extensive (overall ∼5 µs) Molecular Dynamics (MD) simulations of the human VDAC isoforms to detect structural and conformational variations among them, possibly related to specific functional roles of these proteins. Secondary structure analysis of the N-terminal domain shows a high similarity among the three human isoforms of VDAC but with a different plasticity. In particular, the N-terminal domain of the hVDAC1 is characterized by a higher plasticity, with a ∼20% occurrence for the ‘unstructured’ conformation throughout the folded segment, while hVDAC2, containing a peculiar extension of 11 amino acids at the N-terminal end, presents an additional 310-helical folded portion comprising residues 10′ to 3, adhering to the barrel wall. The N-terminal sequences of hVDAC isoforms are predicted to have a low flexibility, with possible consequences in the dynamics of the human VDACs. Clear differences were found between hVDAC1 and hVDAC3 against hVDAC2: a significantly modified dynamics with possible important consequence on the voltage-gating mechanism. Charge distribution inside and at the mouth of the pore is responsible for a different preferential localization of ions with opposite charge and provide a valuable rationale for hVDAC1 and hVDAC3 having a Cl−/K+ selectivity ratio of 1.8, whereas hVDAC2 of 1.4. Our conclusion is that hVDAC isoforms, despite sharing a similar scaffold, have modified working features and a biological work is now requested to give evidence to the described dissimilarities.

电压依赖性阴离子选择通道（Voltage Dependent Anion-selective Channels, VDACs）是定位于线粒体外膜的孔形成蛋白。它们负责介导离子与能量代谢物进入内膜转运系统。哺乳动物体内存在三种VDAC亚型，但其具体功能尚未明确。本研究针对人类VDAC亚型开展了总时长约5 μs的大规模分子动力学（Molecular Dynamics, MD）模拟，以探究三者之间的结构与构象差异，这些差异可能与这些蛋白的特定功能相关。对N端结构域的二级结构分析显示，三种人类VDAC亚型的二级结构高度相似，但结构可塑性存在差异。具体而言，人类VDAC1（hVDAC1）的N端结构域具有更高的可塑性，在整个折叠区段中，“无规构象”的出现比例约为20%；而人类VDAC2（hVDAC2）的N端存在一段特殊的11个氨基酸延伸序列，其包含10′至3位残基的区域可形成额外的3₁₀螺旋结构，并紧贴β桶壁。预测显示，人类VDAC各亚型的N端序列柔韧性较低，这可能会影响人类VDAC的动力学特性。研究发现人类VDAC1、VDAC3与VDAC2之间存在显著差异：其动力学特性发生明显改变，这可能对电压门控机制产生重要影响。孔道内部及孔口的电荷分布决定了带相反电荷离子的不同优先定位方式，这也为下述现象提供了合理的解释：人类VDAC1和VDAC3的Cl−/K+选择性比值为1.8，而VDAC2的该比值为1.4。本研究的结论是，尽管人类VDAC亚型共享相似的支架结构，但其功能特性存在差异，后续需开展生物学实验以验证本研究描述的这些差异。