Kovacević Z, Bajin K
Biochim Biophys Acta. 1982 May 7;687(2):291-5. doi: 10.1016/0005-2736(82)90557-0.
Glutamine transport across the inner membrane of rat liver mitochondria was studied by the method of loading the organelles with [14C]glutamine and by measuring efflux of the metabolite at 0 degree C. The release of [14C]glutamine from loaded mitochondria was prevented by mersalyl, whereas the efflux was started by the addition of glutathione. The rate of glutamine efflux from the mitochondria was measured by the inhibitor stop technique with mersalyl plus N-ethylmaleimide. It was found that up to 10 mM glutamine there is no significant activity of glutaminase, whereas at about 20 mM of the substrate the enzyme is activated. The rate of the efflux measured after the addition of the optimal amount of glutathione was 10 nmol glutamine/min per mg protein. This is 5-times faster than the rate of glutaminase activity at 0 degree C. The pH optimum of glutamine carrier is between 6.5 and 7.0. Low concentration of succinate inhibits the efflux due to formation of pH gradient in coupled mitochondria, whereas a higher concentration of succinate inhibits the carrier directly. 2-Oxoglutarate and glutamate strongly inhibit the rate of glutamine efflux, the inhibition by glutamate being very pronounced at its physiological concentration. D-Glutamine does not inhibit the rate of the efflux, indicating that the transport of L-glutamine is stereospecific.
采用用[14C]谷氨酰胺装载细胞器并在0℃下测量代谢物流出的方法，研究了谷氨酰胺跨大鼠肝脏线粒体内膜的转运。汞撒利可阻止加载的线粒体释放[14C]谷氨酰胺，而加入谷胱甘肽可启动流出。用汞撒利加N-乙基马来酰亚胺的抑制剂停止技术测量线粒体中谷氨酰胺的流出速率。发现高达10 mM的谷氨酰胺时谷氨酰胺酶无明显活性，而在约20 mM的底物时该酶被激活。加入最佳量谷胱甘肽后测得的流出速率为每毫克蛋白质每分钟10 nmol谷氨酰胺。这比0℃时谷氨酰胺酶活性的速率快5倍。谷氨酰胺载体的最适pH在6.5至7.0之间。低浓度的琥珀酸由于在偶联的线粒体中形成pH梯度而抑制流出，而较高浓度的琥珀酸直接抑制载体。2-氧代戊二酸和谷氨酸强烈抑制谷氨酰胺流出速率，谷氨酸在其生理浓度下的抑制作用非常明显。D-谷氨酰胺不抑制流出速率，表明L-谷氨酰胺的转运具有立体特异性。
