Peroxidasin mediates bromination of tyrosine residues in the extracellular matrix
Peroxidasin mediates bromination of tyrosine residues in the extracellular matrix
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DOI：
10.1074/jbc.ra120.014504
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发表时间：
2020-09-04
影响因子：
4.8
通讯作者：
Winterbourn, Christine C.
中科院分区：
文献类型：
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作者：
Bathish, Boushra;Paumann-Page, Martina;Winterbourn, Christine C.
过氧化物酶是一种血红素过氧化物酶，可将溴化物氧化为次溴酸 (HOBr)，次溴酸是一种强氧化剂，可促进基底膜 IV 型胶原蛋白中硫亚胺交联的形成。我们研究了过氧化物酶释放的 HOBr 是否会导致表达过氧化物酶的 PFHR9 细胞中蛋白质的其他氧化修饰，特别是酪氨酸残基的溴化。使用稳定同位素稀释 LC-MS/MS，我们检测到 3-溴酪氨酸的形成，这是 HOBr 介导的蛋白质修饰的特异性生物标志物。正常培养细胞的细胞外基质蛋白中 3-溴酪氨酸的水平为 1.1 mmol/mol 酪氨酸，并且在过氧化物酶抑制剂间苯三酚存在下显着降低。在过氧化物酶敲除细胞中检测到微量的 3-溴酪氨酸。当向嵌入的过氧化酶提供过氧化氢和溴化物时，3-溴酪氨酸在培养物中的细胞生长过程中以及在分离的脱细胞细胞外基质中形成。细胞外基质中 3-溴酪氨酸的水平显着高于细胞内蛋白质，尽管细胞内检测到的量较低。 3-溴酪氨酸水平随着溴化物浓度的升高而升高，而在生理浓度的硫氰酸盐和尿酸盐存在时降低。然而，这些过氧化物酶底物对 IV 型胶原交联显示出中度至最小程度的抑制。我们的研究结果证明过氧化物酶可促进蛋白质中 3-溴酪氨酸的形成。他们表明，过氧化物酶产生的 HOBr 对 IV 型胶原蛋白的交联具有选择性，但不限于此。根据我们的研究结果，使用 3-溴酪氨酸作为 HOBr 氧化损伤的特异性生物标志物值得对与高过氧化物酶表达相关的临床状况进行进一步研究。
Peroxidasin is a heme peroxidase that oxidizes bromide to hypobromous acid (HOBr), a powerful oxidant that promotes the formation of the sulfilimine crosslink in collagen IV in basement membranes. We investigated whether HOBr released by peroxidasin leads to other oxidative modifications of proteins, particularly bromination of tyrosine residues, in peroxidasin-expressing PFHR9 cells. Using stable isotope dilution LC-MS/MS, we detected the formation of 3-bromotyrosine, a specific biomarker of HOBr-mediated protein modification. The level of 3-bromotyrosine in extracellular matrix proteins from normally cultured cells was 1.1 mmol/mol tyrosine and decreased significantly in the presence of the peroxidasin inhibitor, phloroglucinol. A negligible amount of 3-bromotyrosine was detected in peroxidasin-knockout cells. 3-Bromotyrosine formed both during cell growth in culture and in the isolated decellularized extracellular matrix when embedded peroxidasin was supplied with hydrogen peroxide and bromide. The level of 3-bromotyrosine was significantly higher in extracellular matrix than intracellular proteins, although a low amount was detected intracellularly. 3-Bromotyrosine levels increased with higher bromide concentrations and decreased in the presence of physiological concentrations of thiocyanate and urate. However, these peroxidase substrates showed moderate to minimal inhibition of collagen IV crosslinking. Our findings provide evidence that peroxidasin promotes the formation of 3-bromotyrosine in proteins. They show that HOBr produced by peroxidasin is selective for, but not limited to, the crosslinking of collagen IV. Based on our findings, the use of 3-bromotyrosine as a specific biomarker of oxidative damage by HOBr warrants further investigation in clinical conditions linked to high peroxidasin expression.