Wang Qingping, Dubé Daniel, Friesen Richard W, LeRiche Tammy G, Bateman Kevin P, Trimble Laird, Sanghara Joe, Pollex Rebecca, Ramachandran Chidambaram, Gresser Michael J, Huang Zheng
Department of Biochemistry and Molecular Biology, Merck Frosst Centre for Therapeutic Research, Post Office Box 1005, Pointe Claire, Dorval, Quebec H9R 4P8, Canada.
Biochemistry. 2004 Apr 13;43(14):4294-303. doi: 10.1021/bi035986e.
Polyaromatic quinones, such as the environmental pollutants 9,10-phenanthrenediones, elicit a wide range of responses including growth inhibition, immune suppression, and glucose normalization in diabetic models. Yet the molecular mechanisms behind these effects remain controversial. Here we report that many of them are oxygen-dependent and catalytic inactivators of protein tyrosine phosphatases (PTP). Under aerobic conditions, the PTP inactivation by 2-nitro-9,10-phenanthrenedione followed a pseudo-first-order process, with the rate of inactivation increasing nearly linearly with increasing inhibitor concentration, yielding apparent inactivation rate constants of 4300, 387, and 5200 M(-1) s(-1) at pH 7.2 against CD45, PTP1B, and LAR, respectively. The rate of CD45 inactivation increased approximately 25-fold from pH 6.0 to 7.5, with complete inactivation achieved using a catalytic amount (0.05 molar equiv) of the inhibitor. The quinone-catalyzed CD45 inactivation was prevented by catalase or superoxide dismutase. Inactivated CD45 after (125)I-9,10-phenanthrenedione treatment carried no radioactivity, indicating the absence of a stable inhibitor/enzyme complex. The activity of inactivated CD45 was partially restored ( approximately 10%) by hydroxylamine or dithiothreitol, supporting the presence of a small population of sulfenic acid or sulfenyl-amide species. Treatment of PTP1B with 2-nitro-9,10-phenanthrenedione resulted in the specific and sequential oxidation of the catalytic cysteine to the sulfinic and sulfonic acid. These results suggest that reactive oxygen species and the semiquinone radical, continuously generated during quinone-catalyzed redox cycling, mediate the specific catalytic cysteine oxidation. Naturally occurring quinones may act as efficient regulators of protein tyrosine phosphorylation in biological systems. Aberrant phosphotyrosine homeostasis resulting from continued polyaromatic hydrocarbon quinone exposure may play a significant role in their disease etiology.
多环芳烃醌类，如环境污染物9,10 - 菲醌，在糖尿病模型中可引发多种反应，包括生长抑制、免疫抑制和血糖正常化。然而，这些效应背后的分子机制仍存在争议。在此我们报告，其中许多是氧依赖性的蛋白质酪氨酸磷酸酶（PTP）催化失活剂。在有氧条件下，2 - 硝基 - 9,10 - 菲醌使PTP失活遵循准一级反应过程，失活速率随抑制剂浓度增加几乎呈线性增加，在pH 7.2时，针对CD45、PTP1B和LAR的表观失活速率常数分别为4300、387和5200 M⁻¹ s⁻¹。CD45的失活速率从pH 6.0到7.5增加约25倍，使用催化量（0.05摩尔当量）的抑制剂可实现完全失活。过氧化氢酶或超氧化物歧化酶可阻止醌催化的CD45失活。用¹²⁵I - 9,10 - 菲醌处理后失活的CD45不携带放射性，表明不存在稳定的抑制剂/酶复合物。用羟胺或二硫苏糖醇可使失活的CD45活性部分恢复（约10%），这支持存在少量亚磺酸或亚磺酰胺类物质。用2 - 硝基 - 9,10 - 菲醌处理PTP1B导致催化半胱氨酸特异性且顺序性地氧化为亚磺酸和磺酸。这些结果表明，在醌催化的氧化还原循环过程中持续产生的活性氧物种和半醌自由基介导了催化半胱氨酸的特异性氧化。天然存在的醌类可能在生物系统中作为蛋白质酪氨酸磷酸化的有效调节剂。持续暴露于多环芳烃醌导致的异常磷酸酪氨酸稳态可能在其疾病病因中起重要作用。
