Hamasaki N, Matsuyama H, Hirota-Chigita C
Eur J Biochem. 1983 May 16;132(3):531-6. doi: 10.1111/j.1432-1033.1983.tb07394.x.
Phosphoenolpyruvate was found to be transported across the erythrocyte membrane by a carrier-mediated transport system. The transport of phosphoenolpyruvate was competitively inhibited by inorganic phosphate (Ki = 24 mM) and pyridoxal 5-phosphate (Ki = 0.2 mM), whereas the transport was noncompetitively inhibited by L(+)-lactate (Ki = 37 mM). Specific inhibitors for the inorganic anion transport system such as 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid and 4,4'-dinitrostilbene-2,2'-disulfonic acid strongly inhibited the phosphoenolpyruvate transport. The transport was irreversibly inhibited by treating erythrocytes with pyridoxal 5-phosphate and NaBH4. Transport activities of phosphoenolpyruvate and inorganic phosphate in the treated cells were similarly inhibited by pyridoxal 5-phosphate depending on its concentrations. 4,4'-Dinitrostilbene-2,2'-disulfonic acid protected both transport activities against the pyridoxal 5-phosphate/NaBH4 treatment. The major integral membrane protein, band 3, was preferentially labelled by treating erythrocytes with pyridoxal 5-phosphate and NaB[3H]H4. The radioactive incorporation into band 3 was confirmed by two-dimensional gel electrophoresis combining isoelectric focusing in the first dimension and sodium dodecyl sulfate/polyacrylamide gel electrophoresis in the second dimension. Taken together, these results suggest that band 3 mediates the transport of phosphoenolpyruvate as well as inorganic phosphate.
磷酸烯醇丙酮酸通过载体介导的转运系统跨红细胞膜转运。磷酸烯醇丙酮酸的转运受到无机磷酸盐（Ki = 24 mM）和磷酸吡哆醛（Ki = 0.2 mM）的竞争性抑制，而L(+)-乳酸（Ki = 37 mM）对其转运的抑制作用则为非竞争性。无机阴离子转运系统的特异性抑制剂，如4,4'-二异硫氰基芪-2,2'-二磺酸和4,4'-二硝基芪-2,2'-二磺酸，强烈抑制磷酸烯醇丙酮酸的转运。用磷酸吡哆醛和硼氢化钠处理红细胞后，该转运受到不可逆抑制。根据磷酸吡哆醛的浓度，处理后细胞中磷酸烯醇丙酮酸和无机磷酸盐的转运活性同样受到抑制。4,4'-二硝基芪-2,2'-二磺酸可保护两种转运活性免受磷酸吡哆醛/硼氢化钠处理的影响。用磷酸吡哆醛和硼氢化钠[3H]处理红细胞后，主要的整合膜蛋白带3被优先标记。通过二维凝胶电泳（第一维为等电聚焦，第二维为十二烷基硫酸钠/聚丙烯酰胺凝胶电泳）证实了带3中的放射性掺入。综上所述，这些结果表明带3介导了磷酸烯醇丙酮酸以及无机磷酸盐的转运。
