Department of Chemistry - BMC, Uppsala University, Box 576, 751 23 Uppsala, Sweden.
Department of Chemistry, Physical & Theoretical Chemistry Laboratory, University of Oxford, South Parks Road, Oxford OX1 3QZ, UK.
Cell Chem Biol. 2018 Mar 15;25(3):309-317.e4. doi: 10.1016/j.chembiol.2017.12.012. Epub 2018 Jan 18.
The interactions between proteins and biological membranes are important for drug development, but remain notoriously refractory to structural investigation. We combine non-denaturing mass spectrometry (MS) with molecular dynamics (MD) simulations to unravel the connections among co-factor, lipid, and inhibitor binding in the peripheral membrane protein dihydroorotate dehydrogenase (DHODH), a key anticancer target. Interrogation of intact DHODH complexes by MS reveals that phospholipids bind via their charged head groups at a limited number of sites, while binding of the inhibitor brequinar involves simultaneous association with detergent molecules. MD simulations show that lipids support flexible segments in the membrane-binding domain and position the inhibitor and electron acceptor-binding site away from the membrane surface, similar to the electron acceptor-binding site in respiratory chain complex I. By complementing MS with MD simulations, we demonstrate how a peripheral membrane protein uses lipids to modulate its structure in a similar manner as integral membrane proteins.
蛋白质与生物膜之间的相互作用对药物开发很重要，但结构研究却一直难以进行。我们将非变性质谱（MS）与分子动力学（MD）模拟相结合，以揭示辅因子、脂质和抑制剂在周质膜蛋白二氢乳清酸脱氢酶（DHODH）中的结合之间的联系，DHODH 是一种关键的抗癌靶标。通过 MS 对完整的 DHODH 复合物进行检测，发现磷脂通过其带电头基在有限数量的位点结合，而抑制剂布雷奎纳的结合则涉及与去污剂分子的同时结合。MD 模拟表明，脂质支持膜结合域中的柔性片段，并将抑制剂和电子受体结合位点定位在远离膜表面的位置，类似于呼吸链复合物 I 中的电子受体结合位点。通过将 MS 与 MD 模拟相结合，我们展示了外周膜蛋白如何以类似于整合膜蛋白的方式利用脂质来调节其结构。
