Loop 5 region is important for the activity of the long-chain base transporter Rsb1

H. Makuta, K. Obara, A. Kihara
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引用次数: 4

Abstract

Intracellular lipid amounts are regulated not only by metabolism but also by efflux. Yeast Rsb1 is the only known transporter/floppase of the sphingolipid components long-chain bases (LCBs). However, even fundamental knowledge about Rsb1, such as important amino acid residues for activity and substrate recognition, still remains unclear. Rsb1 belongs to the Rta1-like family. To date, it has not been determined whether all family members share a common ability to export LCBs. Here, we revealed that within the Rta1-like family, only Rsb1 suppressed the hypersensitivity of the mutant cells lacking LCB 1-phoshate-degrading enzymes, suggesting that LCB-exporting activity is specific to Rsb1. Rsb1 contains a characteristic region (loop 5), which does not exist in other proteins of the Rta1-like family. We found that deletion of this region caused loss of Rsb1 function. Further mutational analysis of loop 5 revealed that the charged amino acid residues E223, D225 and R236 were important for Rsb1 activity. In addition to LCBs, Rsb1 facilitated the export of 1-hexadecanol, but not palmitic acid, which suggests that Rsb1 recognizes the C1 hydroxyl group. Thus, our findings provide an important clue for understanding the molecular mechanism of LCB export.
环5区对长链碱基转运体Rsb1的活性起重要作用
细胞内脂质量不仅受代谢调节,还受外排调节。酵母Rsb1是鞘脂成分长链碱基(LCBs)唯一已知的转运蛋白/floppase。然而,即使是关于Rsb1的基本知识,如活性和底物识别的重要氨基酸残基,仍然不清楚。rb1属于类rta1家族。迄今为止,尚未确定是否所有家族成员都具有输出lcb的共同能力。在这里,我们发现在rta1样家族中,只有Rsb1抑制缺乏LCB- 1磷酸盐降解酶的突变细胞的超敏反应,这表明LCB输出活性是Rsb1特异性的。rta1含有一个特征区(环5),这在rta1样家族的其他蛋白中不存在。我们发现这个区域的缺失导致了Rsb1功能的丧失。进一步的突变分析表明,环5的带电氨基酸残基E223、D225和R236对rb1活性很重要。除了lcb外,Rsb1还促进了1-十六醇的输出,但不促进棕榈酸的输出,这表明Rsb1识别C1羟基。因此,我们的研究结果为理解LCB输出的分子机制提供了重要线索。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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