School of Pharmacy, Sichuan Industrial Institute of Antibiotics, Chengdu University, Chengdu, China.
Key Laboratory of Birth Defects and Related Disease of Women and Children of MOE, State Key Laboratory of Biotherapy, Department of Obstetrics, West China Second Hospital, Sichuan University, Chengdu, China.
Phys Chem Chem Phys. 2021 Nov 17;23(44):25401-25413. doi: 10.1039/d1cp03756k.
Nucleosides are important precursors of nucleotide synthesis in cells, and nucleoside transporters play an important role in many physiological processes by mediating transmembrane transport and absorption. During nucleoside transport, such proteins undergo a significant conformational transition between the outward- and inward-facing states, which leads to alternating access of the substrate-binding site to either side of the membrane. In this work, a variety of molecular simulation methods have been applied to comparatively investigate the motion modes of human concentrative nucleoside transporter 3 (hCNT3) in three states, as well as global and local cavity conformational changes; and finally, a possible elevator-like transport mechanism consistent with experimental data was proposed. The results of the Gaussian network model (GNM) and anisotropic network model (ANM) show that hCNT3 as a whole tends to contract inwards and shift towards a membrane inside, exhibiting an allosteric process that is more energetically favorable than the rigid conversion. To reveal the complete allosteric process of hCNT3 in detail, a series of intermediate conformations were obtained by an adaptive anisotropic network model (aANM). One of the simulated intermediate states is similar to that of a crystal structure, which indicates that the allosteric process is reliable; the state with lower energy is slightly inclined to the inward-facing structure rather than the expected intermediate crystal structure. The final HOLE analysis showed that except for the outward-facing state, the transport channels were gradually enlarged, which was conductive to the directional transport of nucleosides. Our work provides a theoretical basis for the multistep elevator-like transportation mechanism of nucleosides, which helps to further understand the dynamic recognition between nucleoside substrates and hCNT3 as well as the design of nucleoside anticancer drugs.
核苷是细胞内核苷酸合成的重要前体，核苷转运蛋白通过介导跨膜运输和吸收，在许多生理过程中发挥着重要作用。在核苷转运过程中，这些蛋白质在外向和内向构象之间发生显著的构象转变，导致底物结合位点交替进入膜的两侧。在这项工作中，应用了多种分子模拟方法来比较研究人高亲和性核苷转运蛋白 3（hCNT3）在三种状态下的运动模式，以及整体和局部腔构象变化；最后，提出了一种与实验数据一致的可能的电梯式转运机制。高斯网络模型（GNM）和各向异性网络模型（ANM）的结果表明，hCNT3 整体趋于向内收缩，并向膜内移动，表现出一种变构过程，这种过程比刚性转换更具能量优势。为了详细揭示 hCNT3 的完整变构过程，通过自适应各向异性网络模型（aANM）获得了一系列中间构象。模拟的中间构象之一类似于晶体结构，这表明变构过程是可靠的；能量较低的状态稍微倾向于内向构象，而不是预期的中间晶体结构。最后的 HOLE 分析表明，除了外向构象外，转运通道逐渐扩大，有利于核苷的定向转运。我们的工作为核苷的多步电梯式转运机制提供了理论基础，有助于进一步理解核苷底物与 hCNT3 之间的动态识别以及核苷抗癌药物的设计。
