Catalase is a major antioxidant enzyme located in plant peroxisomes that catalyzes the decomposition of H2O2. Based on our previous transcriptomic (RNA-Seq) and proteomic (iTRAQ) data at different stages of pepper (Capsicum annuum L.) fruit ripening and after exposure to nitric oxide (NO) enriched atmosphere, a broad analysis has allowed us to characterize the functioning of this enzyme. Three genes were identified, and their expression was differentially modulated during ripening and by NO gas treatment. A dissimilar behavior was observed in the protein expression of the encoded protein catalases (CaCat1–CaCat3). Total catalase activity was down-regulated by 50% in ripe (red) fruits concerning immature green fruits. This was corroborated by non-denaturing polyacrylamide gel electrophoresis, where only a single catalase isozyme was identified. In vitro analyses of the recombinant CaCat3 protein exposed to peroxynitrite (ONOO−) confirmed, by immunoblot assay, that catalase underwent a nitration process. Mass spectrometric analysis identified that Tyr348 and Tyr360 were nitrated by ONOO−, occurring near the active center of catalase. The data indicate the complex regulation at gene and protein levels of catalase during the ripening of pepper fruits, with activity significantly down-regulated in ripe fruits. Nitration seems to play a key role in this down-regulation, favoring an increase in H2O2 content during ripening. This pattern can be reversed by the exogenous NO application. While plant catalases are generally reported to be tetrameric, the analysis of the protein structure supports that pepper catalase has a favored quaternary homodimer nature. Taken together, data show that pepper catalase is down-regulated during fruit ripening, becoming a target of tyrosine nitration, which provokes its inhibition.

中文翻译：

---

胡椒过氧化氢酶：对其在果实成熟过程中和一氧化氮调节的广泛分析


过氧化氢酶是位于植物过氧化物酶体中的主要抗氧化酶，可催化 H2O2 的分解。根据我们之前在辣椒 (Capsicum annuum L.) 果实成熟的不同阶段以及暴露于富含一氧化氮 (NO) 的大气后的转录组 (RNA-Seq) 和蛋白质组 (iTRAQ) 数据，广泛的分析使我们能够表征这种酶的功能。鉴定出三个基因，它们的表达在成熟过程中和NO气体处理下受到差异调节。在编码的蛋白质过氧化氢酶 (CaCat1–CaCat3) 的蛋白质表达中观察到不同的行为。与未成熟的绿色水果相比，成熟（红色）水果的总过氧化氢酶活性下调了 50%。非变性聚丙烯酰胺凝胶电泳证实了这一点，其中仅鉴定出单一过氧化氢酶同工酶。对暴露于过氧亚硝酸盐 (ONOO−) 的重组 CaCat3 蛋白进行体外分析，通过免疫印迹测定证实过氧化氢酶经历了硝化过程。质谱分析发现 Tyr348 和 Tyr360 被 ONOO− 硝化，发生在过氧化氢酶活性中心附近。数据表明，在辣椒果实成熟过程中，过氧化氢酶的基因和蛋白质水平存在复杂的调控，成熟果实中的活性显着下调。硝化似乎在这种下调中发挥着关键作用，有利于成熟过程中 H2O2 含量的增加。这种模式可以通过外源一氧化氮的应用来逆转。虽然植物过氧化氢酶通常被报道为四聚体，但蛋白质结构的分析支持辣椒过氧化氢酶具有有利的四元同二聚体性质。 综上所述，数据表明，辣椒过氧化氢酶在果实成熟过程中下调，成为酪氨酸硝化的目标，从而引发其抑制。