Department of Pharmaco-Biology, University of Calabria, Arcavacata di Rende, Cosenza, Italy.
FEBS J. 2010 Mar;277(5):1172-81. doi: 10.1111/j.1742-4658.2009.07550.x. Epub 2010 Jan 27.
The mitochondrial carriers are a family of transport proteins that shuttle metabolites, nucleotides and cofactors across the inner mitochondrial membrane. The genome of Drosophila melanogaster encodes at least 46 members of this family. Only five of these have been characterized, whereas the transport functions of the remainder cannot be assessed with certainty. In the present study, we report the functional identification of two D. melanogaster genes distantly related to the human and yeast thiamine pyrophosphate carrier (TPC) genes as well as the corresponding expression pattern throughout development. Furthermore, the functional characterization of the D. melanogaster mitochondrial thiamine pyrophosphate carrier protein (DmTpc1p) is described. DmTpc1p was over-expressed in bacteria, the purified protein was reconstituted into liposomes, and its transport properties and kinetic parameters were characterized. Reconstituted DmTpc1p transports thiamine pyrophosphate and, to a lesser extent, pyrophosphate, ADP, ATP and other nucleotides. The expression of DmTpc1p in Saccharomyces cerevisiaeTPC1 null mutant abolishes the growth defect on fermentable carbon sources. The main role of DmTpc1p is to import thiamine pyrophosphate into mitochondria by exchange with intramitochondrial ATP and/or ADP.
线粒体载体是一类穿梭代谢物、核苷酸和辅助因子穿过线粒体内膜的转运蛋白。果蝇的基因组至少编码了 46 个该家族的成员。其中只有 5 个得到了鉴定，而其余的转运功能则不能被确定。在本研究中，我们报告了两个与人类和酵母硫胺素焦磷酸载体（TPC）基因在进化上远缘的果蝇基因的功能鉴定，以及它们在整个发育过程中的表达模式。此外，还描述了果蝇线粒体硫胺素焦磷酸载体蛋白（DmTpc1p）的功能特征。DmTpc1p 在细菌中过表达，纯化的蛋白质被重构成脂质体，并对其转运特性和动力学参数进行了表征。重构成的 DmTpc1p 可转运硫胺素焦磷酸，并在一定程度上转运焦磷酸、ADP、ATP 和其他核苷酸。在酿酒酵母 TPC1 缺失突变体中表达 DmTpc1p 可消除其在可发酵碳源上的生长缺陷。DmTpc1p 的主要作用是通过与线粒体内的 ATP 和/或 ADP 交换将硫胺素焦磷酸导入线粒体。
