Molecular Insights into the Binding Specificity of Branched-Chain Amino Acids to Sestrin2: An Integrated Computational Study Using Molecular Docking, MD Simulations, and MMGBSA Analysis

Sestrin2 (SESN2) is a highly conserved stress-inducible protein that serves as a central hub for integrating cellular responses to nutrient availability, oxidative stress, and endoplasmic reticulum (ER) stress. A key function of SESN2 is its role as a direct sensor for the branched-chain amino acid (BCAA) leucine, which modulates the activity of the mechanistic target of rapamycin complex 1 (mTORC1), a master regulator of cell growth and metabolism. While the functional link between leucine and SESN2 is well-established, the precise molecular determinants that confer its high specificity for leucine over other BCAAs, such as isoleucine and valine, remain poorly understood. This study employs an integrated, multi-scale computational approach, combining molecular docking, extensive all-atom molecular dynamics (MD) simulations, and binding free energy calculations, to elucidate the structural and dynamic basis of BCAA-SESN2 recognition. Our thermodynamic analysis reveals a distinct binding affinity hierarchy (Leucine > Isoleucine > Valine), which is primarily driven by superior van der Waals interactions and shape complementarity of leucine's isobutyl side chain within the protein's hydrophobic pocket. Structural and dynamic analyses demonstrate that leucine binding induces a profound allosteric stabilization, causing a global compaction of the protein and establishing a network of long-range correlated motions. This "conformational locking" mechanism funnels the intrinsically flexible apo-protein into a singular, stable, low-energy state. These findings provide a comprehensive, atom-to-global scale model explaining SESN2's function as a specific leucine sensor. Furthermore, by contextualizing these results within the broader landscape of SESN2's mTORC1-independent functions and recent clinical trial outcomes for SESN2 modulators, this work offers a critical framework for understanding its complex role in metabolic diseases, cancer, and neurodegeneration, and provides a rationale for the design of next-generation allosteric therapeutics.Sestrin2（SESN2）是一类高度保守的应激诱导蛋白，作为核心枢纽整合细胞对营养供给、氧化应激及内质网（Endoplasmic Reticulum, ER）应激的应答过程。其关键功能之一是作为支链氨基酸（branched-chain amino acid, BCAA）亮氨酸的直接感受器，可调控雷帕霉素机制性靶点复合物1（mTORC1）的活性——后者是细胞生长与代谢的核心调控因子。尽管亮氨酸与SESN2之间的功能关联已得到充分证实，但其对亮氨酸相较于其他支链氨基酸（如异亮氨酸与缬氨酸）具备高特异性的精确分子决定簇，目前仍知之甚少。本研究采用整合式多尺度计算策略，结合分子对接（molecular docking）、大规模全原子分子动力学（molecular dynamics, MD）模拟与结合自由能计算，以阐明支链氨基酸与SESN2识别过程的结构与动态基础。本研究的热力学分析揭示了明确的结合亲和力层级（亮氨酸＞异亮氨酸＞缬氨酸），该层级主要由亮氨酸的异丁基侧链在蛋白疏水口袋内更优异的范德华相互作用与形状互补性所驱动。结构与动态分析表明，亮氨酸结合可诱导显著的别构稳定效应，引发蛋白整体构象压缩，并构建长程关联运动网络。这种“构象锁定”机制可将固有柔性的脱辅基蛋白引导至单一、稳定的低能构象状态。上述发现构建了一套完整的原子级至全局尺度的模型，阐释了SESN2作为特异性亮氨酸感受器的功能机制。此外，本研究将这些结果置于SESN2不依赖mTORC1的功能范畴以及SESN2调节剂的最新临床试验背景中，为理解其在代谢疾病、癌症与神经退行性疾病中的复杂作用提供了关键框架，并为下一代别构治疗药物的设计提供了理论依据。