Bai Ru-Hsuan, Lin Chun-Chen, Lin Chun-Wei
Department of Chemistry, National Tsing Hua University, Hsinchu, Taiwan 300044.
J Phys Chem Lett. 2025 Jul 3;16(26):6745-6756. doi: 10.1021/acs.jpclett.5c00988. Epub 2025 Jun 24.
The cell membrane is a critical component of cellular architecture, serving not only as a physical barrier enclosing the cytosol but also as a dynamic platform for various biochemical reactions. Due to the unique two-dimensional and fluidic environment of the membrane, reactions that occur on its surface are subject to specific physical constraints. While membrane-mediated reactions are known to play key roles in cellular regulation, their advantages and limitations remain inadequately explored. In this study, we reconstitute a classic proteolytic cleavage reaction at the membrane interface, designed for the real-time kinetic analysis down to the single-molecule level. By systematically altering the enzyme-membrane affinity, we examined enzyme-substrate interactions under various conditions. Our findings reveal that while the membrane environment significantly enhances enzymatic turnover rate, it also imposes diffusion limitations that immediately reduce this turnover rate over time. By adjusting the enzyme's membrane affinity to an intermediate level, we enable the enzyme to "hop" on the membrane surface, overcoming these diffusion constraints and sustaining high enzymatic turnover rate with faster kinetics. These results highlight the dual role of the membrane environment in regulating biochemical reactions, balancing enhanced reactivity with physical limitations. Moreover, the ability to dynamically tune membrane affinity to optimize reactions underscores the cell's capacity to regulate enzymatic processes efficiently. This study provides critical insights into the role of the cell membrane in biochemical reactions and offers a broadly applicable framework for understanding membrane-associated interactions in biological systems.
细胞膜是细胞结构的关键组成部分，不仅作为包围细胞质溶胶的物理屏障，还作为各种生化反应的动态平台。由于膜独特的二维流体环境，发生在其表面的反应受到特定的物理限制。虽然已知膜介导的反应在细胞调节中起关键作用，但其优势和局限性仍未得到充分探索。在本研究中，我们在膜界面重建了一个经典的蛋白水解切割反应，用于单分子水平的实时动力学分析。通过系统地改变酶与膜的亲和力，我们研究了各种条件下的酶-底物相互作用。我们的研究结果表明，虽然膜环境显著提高了酶的周转速率，但它也带来了扩散限制，随着时间的推移会立即降低这种周转速率。通过将酶与膜的亲和力调整到中间水平，我们使酶能够在膜表面“跳跃”，克服这些扩散限制，并以更快的动力学维持高酶周转速率。这些结果突出了膜环境在调节生化反应中的双重作用，在增强反应性与物理限制之间取得平衡。此外，动态调节膜亲和力以优化反应的能力强调了细胞有效调节酶促过程的能力。这项研究为细胞膜在生化反应中的作用提供了关键见解，并为理解生物系统中与膜相关的相互作用提供了一个广泛适用的框架。
