Transport of amino acid-related compounds mediated by L-type amino acid transporter 1 (LAT1): Insights into the mechanisms of substrate recognition
Transport of amino acid-related compounds mediated by L-type amino acid transporter 1 (LAT1): Insights into the mechanisms of substrate recognition
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DOI：
10.1124/mol.61.4.729
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发表时间：
2002-04-01
影响因子：
3.6
通讯作者：
Endou, H
中科院分区：
文献类型：
--
作者：
Uchino, H;Kanai, Y;Endou, H
L型氨基酸转运蛋白1 （LAT1）是一种不依赖Na+的中性氨基酸转运蛋白，服务于氨基酸转运系统L。由于其广泛的底物选择性，L系统被认为是氨基酸相关药物通过质膜渗透的原因。为了了解底物识别的机制，我们利用非洲爪蟾卵母细胞表达系统研究了lat1介导的转运。lat1介导的[C-14]苯丙氨酸摄取被芳香族氨基酸衍生物（包括l -多巴、α -甲基多巴、melphalan、三碘甲状腺原氨酸和甲状腺素）以竞争方式强烈抑制，而苯丙氨酸甲酯、n -甲基苯丙氨酸、多巴胺、酪胺、卡比多巴和droxidopa不抑制[C-14]苯丙氨酸摄取。加巴喷丁，一种γ -氨基酸，也对lat1介导的[C-14]苯丙氨酸摄取具有竞争性抑制作用。虽然大多数抑制LAT1介导摄取的化合物能够诱导预载的[C-14]苯丙氨酸通过强制性交换机制外排到表达LAT1的卵母细胞中，但美法兰、三碘甲状腺原氨酸和甲状腺素没有诱导明显的外排。基于实验和半经验计算分析，提出芳香氨基酸要成为LAT1底物，必须有一个游离羧基和一个氨基。靠近氨基的羰基氧需要计算电荷为-0.55，类似于-0.56，并且不能参与氢键。此外，底物侧链与LAT1底物结合位点之间的疏水相互作用似乎对底物结合至关重要。然而，一旦Connolly可达区域变大和/或分子具有较高的计算logP值（如melphalan、三碘甲状腺原氨酸和甲状腺素），底物就成为阻滞剂。
The L-type amino acid transporter 1 (LAT1) is an Na+ independent neutral amino acid transporter subserving the amino acid transport system L. Because of its broad substrate selectivity, system L has been proposed to be responsible for the permeation of amino acid-related drugs through the plasma membrane. To understand the mechanisms of substrate recognition, we have examined the LAT1-mediated transport using a Xenopus laevis oocyte expression system. LAT1-mediated [C-14] phenylalanine uptake was strongly inhibited in a competitive manner by aromatic-amino acid derivatives including L-dopa, alpha-methyldopa, melphalan, triiodothyronine, and thyroxine, whereas phenylalanine methyl ester, N-methyl phenylalanine, dopamine, tyramine, carbidopa, and droxidopa did not inhibit [C-14] phenylalanine uptake. Gabapentin, a gamma-amino acid, also exerted a competitive inhibition on LAT1-mediated [C-14] phenylalanine uptake. Although most of the compounds that inhibited LAT1-mediated uptake were able to induce the efflux of [C-14] phenylalanine preloaded to the oocytes expressing LAT1 through the obligatory exchange mechanism, melphalan, triiodothyronine, and thyroxine did not induce the significant efflux. Based on the experimental and semiempirical computational analyses, it is proposed that, for an aromatic amino acid to be a LAT1 substrate, it must have a free carboxyl and an amino group. The carbonyl oxygen closer to the amino group needs a computed charge of -0.55similar to -0.56 and must not participate in hydrogen bonding. In addition, the hydrophobic interaction between the substrate side chain and the substrate binding site of LAT1 seems to be crucial for the substrate binding. A substrate, however, becomes a blocker once Connolly accessible areas become large and/or the molecule has a high calculated logP value, such as those for melphalan, triiodothyronine, and thyroxine.