Analysis of the molecular interaction of mitragynine from kratom with human α₁-acid glycoprotein: biophysical and molecular modeling investigations

Mitragynine (MTG), the primary psychoactive alkaloid in Mitragyna speciosa (kratom), has garnered much attention for its therapeutic properties, which is attributed mainly to its selective action on opioid receptors. Despite its clinical potential, the molecular framework of its binding to plasma proteins remains incomplete. Specifically, no studies have thoroughly examined its interaction with α1-acid glycoprotein (AAG), a carrier protein in the circulatory system that influences drug disposition and bioavailability. Hence, this study aims to explore the binding dynamics between MTG and AAG using a combination of spectroscopic, calorimetric, microscopic, and computational methods. Based on isothermal titration calorimetric and fluorescence studies, an intermediate affinity for the MTG–AAG binding was determined (<i>K</i>_a ∼ 10⁵ M⁻¹). Despite evidence of microenvironmental changes around Trp residues, MTG binding did not disrupt the overall structural integrity of AAG. Thermodynamic analysis indicated that the MTG–AAG interaction was energetically favorable, and enthalpy driven mainly by hydrogen bonding and van der Waals forces, with negative entropy change suggesting a more ordered complex formation. Docking analysis showed MTG embedded more deeply within the central cavity of variant F1*S, enhancing complex stability, as opposed to binding near the cavity entrance in variant A. Molecular dynamics simulations supported the stable complexation of MTG with both AAG variants, with variant F1*S maintaining more structural compactness while variant A exhibited slight unfolding upon binding. These findings have clear significance on the potential therapeutic applications of kratom-derived drugs, especially those structurally related to MTG.帽柱木碱（Mitragynine, MTG）是卡痛叶（Mitragyna speciosa, 俗称kratom）中的主要精神活性生物碱，因其对阿片受体的选择性作用而具备显著治疗潜力，因而受到广泛关注。尽管其具备临床应用前景，但目前关于其与血浆蛋白结合的分子机制仍未完全阐明。具体而言，尚无研究深入探讨其与α1-酸性糖蛋白（α1-acid glycoprotein, AAG）的相互作用——该蛋白是循环系统中的载体蛋白，可影响药物的体内处置过程与生物利用度。因此，本研究旨在结合光谱法、量热法、显微成像法与计算方法，探究MTG与AAG之间的结合动力学特征。基于等温滴定量热法与荧光实验结果，本研究确定MTG与AAG的结合亲和力处于中等水平，结合常数K_a约为10^5 M^−1。尽管观察到色氨酸（Trp）残基周围的微环境发生变化，但MTG的结合并未破坏AAG的整体结构完整性。热力学分析表明，MTG与AAG的相互作用在能量上是有利的，且主要由氢键与范德华力驱动的焓变主导，负熵变化则提示复合物形成过程伴随更有序的结构组装。分子对接分析显示，相较于变体A中结合于空腔入口附近，MTG在变体F1*S的中央空腔内嵌入更深，从而提升了复合物的稳定性。分子动力学模拟结果证实，MTG与两种AAG变体均可形成稳定复合物：变体F1*S能够维持更紧密的结构紧凑性，而变体A在结合后则出现轻微的解折叠现象。上述研究结果对于卡痛来源药物（尤其是结构与MTG相关的药物）的潜在治疗应用具有重要指导意义。