Zhang K, Wong K P
Department of Biochemistry, Faculty of Medicine, National University of Singapore, Kent Ridge, Singapore.
Cancer Lett. 1996 Nov 12;108(1):143-51. doi: 10.1016/s0304-3835(96)04457-6.
The formation of the glutathione S-conjugate of monochlorobimane (GSH-bimane) in human colon adenocarcinoma cells was identified by HPLC-fluorimetry and its transport from the cells was found to be temperature-sensitive, saturable and ATP-dependent. The apparent K(m) and Vmax values were 2.4 +/- 0.5 nmol GSH-bimane/10(6) cells and 0.5 +/- 0.1 nmol GSH-bimane/min per 10(6) cells, respectively. This active transport of GSH-bimane was inhibited by low micromolar concentrations of classical uncouplers of oxidative phosphorylation, namely carbonylcyanide p-trifluoromethoxyphenylhydrazone (FCCP), carbonylcyanide m-chlorophenylhydrazone (CCCP) and 2,4-dinitrophenol (DNP). The efflux of GSH-bimane was competitively inhibited by chlorambucil (CMB) and 1-chloro-2,4-dinitrobenzene (CDNB), two other substrates of GST. This study demonstrates the presence and kinetic measurements of the glutathione S-conjugate export (GS-X) pump in human colon cancer cells, an export pump whose function has been implicated in the phenomenon of multidrug resistance.
通过高效液相色谱 - 荧光法鉴定了人结肠腺癌细胞中一氯双硫腙的谷胱甘肽S - 共轭物（谷胱甘肽 - 双硫腙，GSH - bimane）的形成，并且发现其从细胞中的转运对温度敏感、具有饱和性且依赖ATP。表观K(m)和Vmax值分别为2.4±0.5 nmol谷胱甘肽 - 双硫腙/10(6)个细胞和0.5±0.1 nmol谷胱甘肽 - 双硫腙/分钟·10(6)个细胞。谷胱甘肽 - 双硫腙的这种主动转运受到低微摩尔浓度的经典氧化磷酸化解偶联剂的抑制，即羰基氰对 - 三氟甲氧基苯腙（FCCP）、羰基氰间 - 氯苯腙（CCCP）和2,4 - 二硝基苯酚（DNP）。谷胱甘肽 - 双硫腙的外排受到苯丁酸氮芥（CMB）和1 - 氯 - 2,4 - 二硝基苯（CDNB）的竞争性抑制，这两种物质是谷胱甘肽S - 转移酶（GST）的其他底物。本研究证明了人结肠癌细胞中谷胱甘肽S - 共轭物输出（GS - X）泵的存在及其动力学测量，这种输出泵的功能与多药耐药现象有关。
