Identification of Kynoxazine, a Novel Fluorescent Product of the Reaction between 3-Hydroxykynurenine and Erythrulose in the Human Lens, and Its Role in Protein Modification
Identification of Kynoxazine, a Novel Fluorescent Product of the Reaction between 3-Hydroxykynurenine and Erythrulose in the Human Lens, and Its Role in Protein Modification
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DOI：
10.1074/jbc.m116.716621
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发表时间：
2016-04-29
影响因子：
4.8
通讯作者：
Nagaraj, Ram H.
中科院分区：
文献类型：
--
作者：
Rakete, Stefan;Nagaraj, Ram H.
犬尿氨酸途径代谢物和抗坏血酸降解产物存在于人类晶状体中。在这项研究中，我们发现赤藓酮糖（一种主要的抗坏血酸降解产物）会自发地与 3-羟基犬尿氨酸反应形成荧光产物。产物的结构表征表明它是2-氨基-4-(2-羟基-3-(2-羟乙基)-2H-苯并[b][1,4]恶嗪-5-基)-4-氧代丁酸，我们将其命名为kynoxazine。与3-羟基犬尿氨酸不同，3-羟基犬尿氨酸葡萄糖苷和犬尿氨酸不能形成犬恶嗪样化合物，这表明3-羟基犬尿氨酸中的氨基酚部分对于犬恶嗪的形成至关重要。这一推理得到了模型化合物 1-(2-氨基-3-羟基苯基)乙烷-1-酮的证实，它是一种缺乏 3-羟基犬尿氨酸氨基酸部分的氨基酚。超高效液相色谱-串联质谱分析表明，犬恶嗪在人晶状体中的含量为 0 至 64 pmol/mg 晶状体。喹恶嗪和赤藓酮糖在生理条件下降解生成 3-脱氧苏酮，其通过形成精氨酸加合物、3-脱氧苏酮衍生的氢咪唑酮和赖氨酸-精氨酸交联加合物、3-脱氧苏酮衍生的氢咪唑亚胺交联来修饰和交联蛋白质。超高效液相色谱-串联质谱定量显示，老化镜片中发生了32-169 pmol/mg蛋白质的3-脱氧苏酮衍生的氢化咪唑啉酮和1.1-11.2 pmol/mg蛋白质的3-脱氧苏酮衍生的氢化咪唑亚胺交联。总而言之，这些结果证明了一种新的生化机制，抗坏血酸氧化和犬尿氨酸途径相互交织，可以促进老化人类晶状体中的蛋白质修饰和交联。
Kynurenine pathway metabolites and ascorbate degradation products are present in human lenses. In this study, we showed that erythrulose, a major ascorbate degradation product, reacts spontaneously with 3-hydroxykynurenine to form a fluorescent product. Structural characterization of the product revealed it to be 2-amino-4-(2-hydroxy-3-(2-hydroxyethyl)-2H-benzo[b][1,4]oxazin-5-yl)-4-oxobutanoic acid, which we named kynoxazine. Unlike 3-hydroxykynurenine, 3-hydroxykynurenine glucoside and kynurenine were unable to form a kynoxazine-like compound, which suggested that the aminophenol moiety in 3-hydroxykynurenine is essential for the formation of kynoxazine. This reasoning was confirmed using a model compound, 1-(2-amino-3-hydroxyphenyl)ethan-1-one, which is an aminophenol lacking the amino acid moiety of 3-hydroxykynurenine. Ultra-performance liquid chromatography-tandem mass spectrometry analyses showed that kynoxazine is present in the human lens at levels ranging from 0 to 64 pmol/mg lens. Kynoxazine as well as erythrulose degraded under physiological conditions to generate 3-deoxythreosone, which modified and cross-linked proteins through the formation of an arginine adduct, 3-deoxythreosone-derived hydroimidazolone, and a lysine-arginine cross-linking adduct, 3-deoxythreosone-derived hydroimidazolimine cross-link. Ultra-performance liquid chromatography-tandem mass spectrometry quantification showed that 32-169 pmol/mg protein of 3-deoxythreosone-derived hydroimidazolone and 1.1-11.2 pmol/mg protein of 3-deoxythreosone-derived hydroimidazolimine cross-link occurred in aging lenses. Taken together, these results demonstrate a novel biochemical mechanism by which ascorbate oxidation and the kynurenine pathway intertwine, which could promote protein modification and cross-linking in aging human lenses.