Maffia M, Cassano G, Marcucci D, Vilella S, Storelli C
Laboratorio di Fisiologia, Università di Lecce, Italy.
Biochim Biophys Acta. 1990 Aug 10;1027(1):8-16. doi: 10.1016/0005-2736(90)90040-u.
The mechanism of Na+/L-proline cotransport, present on brush-border membrane (BBM) vesicles of the European eel intestine, was studied. Initial cotransport rates, depending on increasing proline and Na+ concentrations in the extravesicular medium (zero-trans conditions), were measured by monitoring the decay of an inside-negative membrane potential, i.e. the fluorescence quenching of the voltage-sensitive cyanine dye 3,3'-diethylthiacarbocyanine iodide (DiS-C2(5)). By simultaneously estimating the substrate-dependent Na(+)-influx (with the fluorescent dye) and the Na(+)-dependent [3H]substrate influx, it was concluded that proline was cotransported with 1 Na+ ion and glucose with 2 Na+ ions. The kinetics of proline/Na+ cotransport were then investigated. Graphical analysis excluded a ping-pong mechanism. Under rapid equilibrium assumptions, by fitting model equations to rate values it was possible to exclude the random and the ordered Na+/proline mechanisms. Therefore, in eel intestinal BBM vesicles, the mechanism of proline/Na+ cotransport is ordered and prolineout binds to the carrier prior to Na+out.
对欧洲鳗鲡肠道刷状缘膜（BBM）囊泡上存在的Na⁺/L-脯氨酸共转运机制进行了研究。通过监测膜内负电位的衰减，即电压敏感的花菁染料3,3'-二乙基硫代碳菁碘化物（DiS-C₂(5)）的荧光猝灭，测定了初始共转运速率，该速率取决于细胞外介质中脯氨酸和Na⁺浓度的增加（零转运条件）。通过同时估计底物依赖性的Na⁺内流（使用荧光染料）和Na⁺依赖性的[³H]底物内流，得出脯氨酸与1个Na⁺离子共转运，葡萄糖与2个Na⁺离子共转运的结论。随后研究了脯氨酸/Na⁺共转运的动力学。图形分析排除了乒乓机制。在快速平衡假设下，通过将模型方程拟合到速率值，可以排除随机和有序的Na⁺/脯氨酸机制。因此，在鳗鲡肠道BBM囊泡中，脯氨酸/Na⁺共转运机制是有序的，并且脯氨酸外排先于Na⁺外排与载体结合。
