Tahiliani A G, Keene T, Kaplan R S
Department of Biomedical Sciences, College of Allied Health, University of South Alabama, Mobile 36688.
J Bioenerg Biomembr. 1992 Dec;24(6):635-40. doi: 10.1007/BF00762356.
The effect of protein labeling agents on coenzyme A (CoA) transport into isolated rat heart mitochondria was studied. CoA transport was substantially inhibited by sulfhydryl reagents (mersalyl, pCMB) as well as by the tyrosine-selective reagent N-acetylimidazole. The effect of pCMB was reversed by DTT. Moreover, CoA uptake was completely abolished by agents selective for lysine and amino terminal residues (pyridoxal 5-phosphate, dansyl chloride). In contrast arginine-selective reagents (2, 3-butanedione, phenylglyoxal) caused considerably less inhibition of CoA uptake. Moreover, partial inhibition of transport was observed with the stilbene disulfonic acid derivatives DIDS and SITS. Finally, measurement of the effects of the labeling agents on the mitochondrial membrane potential indicated that the inhibition of CoA transport into mitochondria is not a secondary effect that arises from an alteration in the electric potential gradient across the inner mitochondrial membrane. These results provide the first information on the types of amino acid residues that may be essential to the CoA transport mechanism and provide additional support for the existence of a CoA transport protein within the mitochondrial inner membrane. Furthermore, the identification of effective inhibitors of the CoA transport system will greatly facilitate the functional reconstitution of this transporter in a proteoliposomal system following its solubilization and purification.
研究了蛋白质标记剂对辅酶A（CoA）转运至离体大鼠心脏线粒体的影响。巯基试剂（汞撒利、对氯汞苯甲酸）以及酪氨酸选择性试剂N - 乙酰咪唑均可显著抑制CoA的转运。二硫苏糖醇可逆转对氯汞苯甲酸的作用。此外，赖氨酸和氨基末端残基选择性试剂（磷酸吡哆醛、丹磺酰氯）可完全消除CoA的摄取。相比之下，精氨酸选择性试剂（2,3 - 丁二酮、苯乙二醛）对CoA摄取的抑制作用要小得多。此外，用芪二磺酸衍生物二碘水杨酸二钠（DIDS）和4 - 乙酰氨基 - 4'- 异硫氰酸芪 - 2,2'- 二磺酸（SITS）观察到转运受到部分抑制。最后，对标记剂对线粒体膜电位影响的测量表明，CoA转运至线粒体的抑制并非是由于线粒体内膜跨膜电位梯度改变而产生的继发效应。这些结果首次提供了有关对CoA转运机制可能至关重要的氨基酸残基类型的信息，并为线粒体内膜中存在CoA转运蛋白提供了额外支持。此外，CoA转运系统有效抑制剂的鉴定将极大地促进该转运体在溶解和纯化后在蛋白脂质体系统中的功能重建。
