Zhang Tao, Lu Haojie, Li Weijun, Hu Ronggui, Chen Zi
Department of Laboratory Medicine, Huashan Hospital, Fudan University, 12 Central Urumqi Road, Shanghai 200040, China.
Shanghai Cancer Center and Key Laboratory of Glycoconjugates Research Ministry of Public Health, Fudan University, Shanghai 200032, China.
Int J Mol Sci. 2015 Nov 10;16(11):26871-9. doi: 10.3390/ijms161125994.
The identification of arsenic direct-binding proteins is essential for determining the mechanism by which arsenic trioxide achieves its chemotherapeutic effects. At least two cysteines close together in the amino acid sequence are crucial to the binding of arsenic and essential to the identification of arsenic-binding proteins. In the present study, arsenic binding proteins were pulled down with streptavidin and identified using a liquid chromatograph-mass spectrometer (LC-MS/MS). More than 40 arsenic-binding proteins were separated, and redox-related proteins, glutathione S-transferase P1 (GSTP1), heat shock 70 kDa protein 9 (HSPA9) and pyruvate kinase M2 (PKM2), were further studied using binding assays in vitro. Notably, PKM2 has a high affinity for arsenic. In contrast to PKM2, GSTP1and HSPA9 did not combine with arsenic directly in vitro. These observations suggest that arsenic-mediated acute promyelocytic leukaemia (APL) suppressive effects involve PKM2. In summary, we identified several arsenic binding proteins in APL cells and investigated the therapeutic mechanisms of arsenic trioxide for APL. Further investigation into specific signal pathways by which PKM2 mediates APL developments may lead to a better understanding of arsenic effects on APL.
鉴定砷直接结合蛋白对于确定三氧化二砷实现其化疗效果的机制至关重要。在氨基酸序列中至少两个紧密相邻的半胱氨酸对于砷的结合至关重要，并且对于鉴定砷结合蛋白必不可少。在本研究中，用链霉亲和素拉下砷结合蛋白，并使用液相色谱 - 质谱联用仪（LC-MS/MS）进行鉴定。分离出40多种砷结合蛋白，并使用体外结合试验进一步研究了氧化还原相关蛋白、谷胱甘肽S-转移酶P1（GSTP1）、热休克70kDa蛋白9（HSPA9）和丙酮酸激酶M2（PKM2）。值得注意的是，PKM2对砷具有高亲和力。与PKM2相反，GSTP1和HSPA9在体外不直接与砷结合。这些观察结果表明，砷介导的急性早幼粒细胞白血病（APL）抑制作用涉及PKM2。总之，我们在APL细胞中鉴定了几种砷结合蛋白，并研究了三氧化二砷对APL的治疗机制。进一步研究PKM2介导APL发展的特定信号通路可能会更好地理解砷对APL的作用。
