Key Laboratory of Functional Small Organic Molecule, Ministry of Education and College of Life Science, Jiangxi Normal University, 99 Ziyang Road, Nanchang, Jiangxi 330022, China.
Key Laboratory of Green Chemistry, Jiangxi Province and College of Chemistry and Chemical Engineering, Jiangxi Normal University, 99 Ziyang Road, Nanchang, Jiangxi, 330022, China.
Clin Chim Acta. 2015 Feb 20;441:163-70. doi: 10.1016/j.cca.2014.12.014. Epub 2014 Dec 17.
It is demonstrated that levels of protein-bound chlorotyrosine, nitrotyrosine and myeloperoxidase (MPO), a protein that catalyzes generation of chlorinating and nitrating oxidants, serve as independent predictors of cardiovascular disease.
Immunoprecipitation and Western blot were used to analyze protein concentration, nitration and chlorination. LC-MS/MS was used to identify nitrated and chlorinated sites of Tyr from immunoprecipitated serum proteins.
Apolipoprotein A-I (apoA-I), the primary protein constituent of high density lipoprotein (HDL), was identified as a selective target for MPO-catalyzed nitration and chlorination in patients with type 2 diabetes. The serum proteins from diabetic subjects showed that the levels of apoA-I nitration and chlorination were clearly increased, whereas apoA-I concentration and cholesterol efflux activity were significantly decreased. MPO as a likely mechanism for oxidative modification of apoA-I in vivo was apparently facilitated by MPO binding to apoA-I. Subsequently, it was found that Tyr 192 was the major nitration and chlorination site in apoA-I from diabetic serum. Further studies in vitro revealed that besides the classic inhibition in cholesterol efflux activities, MPO-catalyzed oxidation could result in a loss of anti-apoptotic activity of lipoprotein.
ApoA-I undergoes MPO-mediated oxidation in serum from diabetic patients compared to non-diabetic subjects and MPO-catalyzed modification may impair the anti-apoptotic properties of HDL in vitro.
研究表明，蛋白质结合的氯酪氨酸、硝基酪氨酸和髓过氧化物酶（MPO）水平可作为心血管疾病的独立预测因子，MPO 是一种催化生成氯化和硝化氧化剂的蛋白质。
采用免疫沉淀和 Western blot 分析蛋白浓度、硝化和氯化。采用 LC-MS/MS 鉴定免疫沉淀血清蛋白中 Tyr 的硝化和氯化位点。
载脂蛋白 A-I（apoA-I）是高密度脂蛋白（HDL）的主要蛋白成分，在 2 型糖尿病患者中被鉴定为 MPO 催化硝化和氯化的选择性靶标。糖尿病患者的血清蛋白显示，apoA-I 的硝化和氯化水平明显增加，而 apoA-I 浓度和胆固醇外排活性显著降低。MPO 作为体内 apoA-I 氧化修饰的可能机制，显然通过 MPO 与 apoA-I 的结合而得到促进。随后发现，apoA-I 中的 Tyr192 是 apoA-I 硝化和氯化的主要位点。进一步的体外研究表明，除了经典的胆固醇外排活性抑制外，MPO 催化的氧化还可能导致脂蛋白抗凋亡活性丧失。
与非糖尿病患者相比，糖尿病患者血清中的 apoA-I 经历了 MPO 介导的氧化，MPO 催化的修饰可能会损害 HDL 的体外抗凋亡特性。
