Zaia Kimberly A, Reimer Richard J
Department of Neurology and Neurological Sciences and Neuroscience Graduate Program, Stanford University, Stanford, California 94305, USA.
J Biol Chem. 2009 Mar 27;284(13):8439-48. doi: 10.1074/jbc.M806407200. Epub 2009 Jan 15.
The SLC6 family of structurally related, Na(+)-dependent transporter proteins is responsible for presynaptic reuptake of the majority of neurotransmitters. Within this family are a number of orphan transporters, including NTT4/XT1 (SLC6A17), a protein first identified over 15 years ago. NTT4/XT1 is expressed exclusively in the nervous system and specifically on synaptic vesicles in glutamatergic and some GABAergic neurons. Despite extensive efforts by a number of groups, no substrate has been reported for NTT4/XT1. Here we use a combination of molecular manipulations to increase expression of the NTT4/XT1 protein at the plasma membrane and to directly demonstrate that it catalyzes neutral amino acid transport. The substrate profile of the NTT4/XT1-dependent activity is similar to that of the closely related B(0)AT2/SBAT1 (SLC6A15), including a submillimolar apparent affinity for proline and leucine and a low millimolar apparent affinity for glutamine. The transport activity is Na(+)-dependent and Cl(-)-independent and is inhibited by low pH as is SLC6A15, suggesting redundant roles for these proteins. This characterization of NTT4/XT1 offers important insights into neurotransmitter metabolism as well as the mechanistic differences among the structurally related, but functionally divergent, SLC6 proteins.
结构相关的SLC6家族是一类依赖钠离子的转运蛋白，负责大多数神经递质的突触前再摄取。该家族中有许多孤儿转运体，包括NTT4/XT1（SLC6A17），一种15年前首次被鉴定的蛋白质。NTT4/XT1仅在神经系统中表达，且特异性地表达于谷氨酸能神经元和一些γ-氨基丁酸能神经元的突触小泡上。尽管多个研究小组付出了巨大努力，但尚未有关于NTT4/XT1底物的报道。在此，我们采用多种分子操作相结合的方法，提高NTT4/XT1蛋白在质膜上的表达，并直接证明它催化中性氨基酸转运。NTT4/XT1依赖活性的底物谱与密切相关的B(0)AT2/SBAT1（SLC6A15）相似，包括对脯氨酸和亮氨酸具有亚毫摩尔级的表观亲和力，对谷氨酰胺具有低毫摩尔级的表观亲和力。其转运活性依赖钠离子且不依赖氯离子，并且像SLC6A15一样受低pH抑制，这表明这些蛋白质具有冗余作用。对NTT4/XT1的这一特性描述为神经递质代谢以及结构相关但功能不同的SLC6蛋白之间的机制差异提供了重要见解。
