Laboratorio de Bioquímica, Departamento de Biología Vegetal, Facultad de Agronomía, Universidad de la República , Av. E. Garzón 780, CP 12900 Montevideo, Uruguay.
J Phys Chem B. 2014 Jan 9;118(1):37-47. doi: 10.1021/jp407773u. Epub 2013 Dec 24.
The accumulation of proline (Pro) and overproduction of reactive oxygen species (ROS) by plants exposed to stress is well-documented. In vitro assays show that enzyme inactivation by hydroxyl radicals ((•)OH) can be avoided in the presence of Pro, suggesting this amino acid might act as a (•)OH scavenger. Although production of hydroxyproline (Hyp) has been hypothesized in connection with such antioxidant activity, no evidence on the detailed mechanism of scavenging has been reported. To elucidate whether and how Hyp might be produced, we used density functional theory calculations coupled to a polarizable continuum model to explore 27 reaction channels including H-abstraction by (•)OH and (•)OH/H2O addition. The structure and energetics of stable species and transition states for each reaction channel were characterized at the PCM-(U)M06/6-31G(d,p) level in aqueous solution. Evidence is found for a main pathway in which Pro scavenges (•)OH by successive H-abstractions (ΔG(‡,298) = 4.1 and 7.5 kcal mol(-1)) to yield 3,4-Δ-Pro. A companion pathway with low barriers yielding Δ(1)-pyrroline-5-carboxylate (P5C) is also supported, linking with 5-Hyp through hydration. However, this connection remains unlikely in stressed plants because P5C would be efficiently recycled to Pro (contributing to its accumulation) by P5C reductase, hypothesis coined here as the "Pro-Pro cycle".
植物在受到胁迫时会积累脯氨酸（Pro）并产生过量的活性氧物种（ROS），这一现象已有充分的文献记载。体外实验表明，在脯氨酸存在的情况下，酶可以避免被羟基自由基（（•）OH）失活，这表明这种氨基酸可能充当（•）OH 清除剂。尽管已经假设羟脯氨酸（Hyp）的产生与这种抗氧化活性有关，但尚未有关于清除详细机制的证据。为了阐明 Hyp 是否以及如何产生，我们使用密度泛函理论计算结合极化连续体模型来探索 27 个反应通道，包括（•）OH 的 H 原子提取和（•）OH/H2O 的加成。在水溶液中，在 PCM-（U）M06/6-31G（d，p）水平上对每个反应通道的稳定物种和过渡态的结构和能量进行了表征。有证据表明，Pro 通过连续的 H 原子提取（ΔG(‡,298)=4.1 和 7.5 kcal mol(-1))来清除（•）OH，从而产生 3,4-Δ-Pro，这是一条主要途径。还支持一条具有低能垒的伴生途径，生成 Δ(1)-吡咯啉-5-羧酸（P5C），通过水合作用与 5-Hyp 相连。然而，由于 P5C 还原酶可将 P5C 有效地循环回 Pro（有助于其积累），因此这种联系在应激植物中不太可能发生，我们在这里提出了“Pro-Pro 循环”的假说。
