Sadongo Victor W, Kim Eojin, Kim Seoyoon, Wijesinghe W C Bhashini, Lee Taeseung, Choi Jeong-Mo, Min Duyoung
Department of Chemistry, Ulsan National Institute of Science and Technology, Ulsan, Republic of Korea.
Department of Chemistry, Pusan National University, Busan, Republic of Korea.
Nat Commun. 2025 Aug 9;16(1):7366. doi: 10.1038/s41467-025-62852-1.
Dimerization of transmembrane (TM) proteins is a fundamental process in cellular membranes, central to numerous physiological and pathological pathways, and increasingly recognized as a promising therapeutic target. Although often described as a simple two-state transition from monomers to dimers, the process following monomer diffusion-referred to as post-diffusion dimerization-is likely more intricate due to complex inter-residue interactions. Here, we present a single-molecule tweezer platform that directly profiles these post-diffusion transitions during TM protein dimerization. This approach captures reversible dimerization events of individual TM dimers, revealing previously hidden intermediate states that emerge after monomer diffusion. By integrating measurements of intermediates, kinetics, and energy landscapes with molecular dynamics simulations, we delineate the dimerization pathway and dissect how residue interactions and lipid bilayers influence the process. Furthermore, our platform allows for the targeted analysis of localized perturbations-such as those induced by peptide binding or site-directed mutagenesis-demonstrating its utility for probing the mechanisms of TM dimer-targeting therapeutics at single-molecule resolution.
跨膜（TM）蛋白的二聚化是细胞膜中的一个基本过程，是众多生理和病理途径的核心，并且越来越被认为是一个有前景的治疗靶点。尽管通常被描述为从单体到二聚体的简单双态转变，但单体扩散后发生的过程——称为扩散后二聚化——由于复杂的残基间相互作用，可能更为复杂。在这里，我们展示了一个单分子镊子平台，该平台可直接描绘TM蛋白二聚化过程中的这些扩散后转变。这种方法捕获单个TM二聚体的可逆二聚化事件，揭示单体扩散后出现的先前隐藏的中间状态。通过将中间体、动力学和能量景观的测量与分子动力学模拟相结合，我们描绘了二聚化途径，并剖析了残基相互作用和脂质双层如何影响这一过程。此外，我们的平台允许对局部扰动进行靶向分析——例如由肽结合或定点诱变诱导的扰动——证明了其在单分子分辨率下探测TM二聚体靶向治疗机制的实用性。
