化学试剂

低氧激活前药(HAP)是靶向肿瘤低氧微环境的重要治疗策略。三甲基锁醌丙酸(Q_3PA)作为一种醌类HAP，可在肿瘤低氧条件下经醌氧化还原酶1(NQO1)还原，触发分子内环化并释放活性药效团，已被广泛用于抗肿瘤前药和荧光探针的设计。然而，其具体的酶催化还原机制尚不明确，限制了进一步的理性优化。采用密度泛函理论(DFT)和分子对接方法，系统探讨了辅酶还原型黄素二核苷酸(FADH₂)介导的Q3PA双电子还原机理及其与NQO1活性中心的相互作用模式。通过构建4条可能的反应路径并计算各驻点的结构与能量，明确了反应的优势路径:首先由FADH₂向Q_3PA的羰基氧原子传递2e~-/1H⁺，生成半醌阴离子中间体;随后由组氨酸残基提供第二个质子，生成对氢醌结构;最后经分子内环化形成内酯，释放药效团。分子对接结果显示，Q_3PA的对苯醌部分以平行方式堆叠于FADH₂异咯嗪环上方，形成稳定的π-π堆积作用;其羰基与Tyr126和Tyr128形成氢键，同时与Pro68、Trp105、Phe106、His161和Phe178等残基产生疏水相互作用，确保了底物在活性中心的准确定位。研究从原子层面揭示了Q_3PA在NQO1催化下的还原路径及关键结构特征，明确了反应优势路径及控速步骤，阐明了底物与酶的识别机制，为设计新型的靶向性抗癌药物提供了理论依据。

Hypoxia-activated prodrugs( HAPs) represent an important therapeutic strategy targeting the hypoxic tumor microenvironment. Trimethyl lock quinone propionic acid( Q₃ PA),a quinone-based HAP,can be reduced by quinone oxidoreductase 1( NQO1) under tumor hypoxia,triggering intramolecular cyclization and the release of active pharmacophores.It has been widely employed in the design of antitumor prodrugs and fluorescent probes. However,the specific enzymatic reduction mechanism of Q₃ PA remains unclear,limiting its further rational optimization. Density functional theory( DFT) and molecular docking methods were employed to systematically investigate the two-electron reduction mechanism of Q₃ PA mediated by the coenzyme reduced flavin adenine dinucleotide( FADH₂) and its interaction mode within the NQO1 active center.By constructing four possible reaction pathways and calculating the structures and energies of each stationary point,the dominant reaction pathway was elucidated: Initially,FADH₂ delivered 2e~-/1H~+to the carbonyl oxygen atom of Q₃ PA,generating a semiquinone anion intermediate.Subsequently,a histidine residue provided a second proton to form the hydroquinone structure.Finally,intramolecular cyclization yielded a lactone,releasing the pharmacophore. Molecular docking results revealed that the benzoquinone moiety of Q₃ PA stacks parallelly above the isoalloxazine ring of FADH₂,forming stable π-π interactions.Its carbonyl groups form hydrogen bonds with Tyr126 and Tyr128,while hydrophobic interactions with residues including Pro68,Trp105,Phe106,His161,and Phe178 further ensure precise substrate positioning within the active center.This study elucidated the reduction pathway and key structural features of Q₃ PA catalyzed by NQO1 at the atomic level,identifies the dominant pathway and rate-determining step,and clarifies the substrate-enzyme recognition mechanism,providing a theoretical basis for the design of novel hypoxia-targeted antitumor drugs.