Department of Physics, Florida International University, Miami, FL, USA.
Department of Physics and Biomolecular Sciences Institute, Florida International University, Miami, FL, USA.
Methods Mol Biol. 2025;2870:315-340. doi: 10.1007/978-1-0716-4213-9_16.
Multi-antimicrobial extrusion (MATE) transporter membrane proteins provide drug and toxin resistivity by expelling compounds from cells. MATE proteins can be pictured as V-shaped. To regulate its functioning, the protein structure can switch between outward-facing (OF) and inward-facing (IF). Pyrococcus furiosus MATE (PfMATE) is the only member of the multidrug/oligosaccharidyl-lipid/polysaccharide (MOP) superfamily that has available both the IF and OF crystal structures. With the availability of both the IF and OF structures, we are able to perform computational investigations to determine how protonation of specific amino acids causes a cascade of changes in the protein conformation that allow PfMATE to change its state from OF to IF in order to regulate its antiporter function. Using a variety of computational and theoretical techniques, we investigated four different systems of IF and OF PfMATE along with the native archaeal lipid bilayer, without or with protonation at the experimentally determined locations within the protein. We performed molecular dynamics (MD) simulations to investigate the flexibility of the four different PfMATE structures and also performed targeted molecular dynamics (TMD) simulations, during which we observed occluded conformations. Our analysis of hydrogen bond changes, potential of mean force, dynamic network analysis, and transfer entropy analysis provides information on how protonation can induce cascading structural changes responsible for the transition between the IF and OF configurations.
多抗菌体挤出（MATE）转运蛋白膜通过将化合物从细胞中排出提供药物和毒素抗性。MATE 蛋白可以被描绘成 V 形。为了调节其功能，蛋白质结构可以在向外（OF）和向内（IF）之间切换。Pyrococcus furiosus MATE（PfMATE）是唯一具有可用 IF 和 OF 晶体结构的多药/寡糖脂/多糖（MOP）超家族成员。由于具有 IF 和 OF 结构，我们能够进行计算研究，以确定特定氨基酸的质子化如何引起蛋白质构象的级联变化，从而使 PfMATE 能够从 OF 状态转变为 IF 状态，以调节其逆向转运功能。使用各种计算和理论技术，我们研究了 IF 和 OF PfMATE 的四个不同系统，以及没有或在蛋白质中实验确定位置质子化的天然古菌脂质双层。我们进行了分子动力学（MD）模拟，以研究四个不同 PfMATE 结构的柔韧性，并且还进行了靶向分子动力学（TMD）模拟，在此期间我们观察到闭塞构象。我们对氢键变化、平均力势能、动态网络分析和传递熵分析的分析提供了关于质子化如何诱导引发 IF 和 OF 构象之间转变的级联结构变化的信息。
