MhsT of Bacillus halodurans is a transporter of hydrophobic amino acids and a homologue of the eukaryotic SLC6 family of Na⁺‐dependent symporters for amino acids, neurotransmitters, osmolytes, or creatine. The broad range of transported amino acids by MhsT prompted the investigation of the substrate recognition mechanism. Here, we report six new substrate‐bound structures of MhsT, which, in conjunction with functional studies, reveal how the flexibility of a Gly‐Met‐Gly (GMG) motif in the unwound region of transmembrane segment 6 (TM6) is central for the recognition of substrates of different size by tailoring the binding site shape and volume. MhsT mutants, harboring substitutions within the unwound GMG loop and substrate binding pocket that mimick the binding sites of eukaryotic SLC6A18/B0AT3 and SLC6A19/B0AT1 transporters of neutral amino acids, exhibited impaired transport of aromatic amino acids that require a large binding site volume. Conservation of a general (G/A/C)ΦG motif among eukaryotic members of SLC6 family suggests a role for this loop in a common mechanism for substrate recognition and translocation by SLC6 transporters of broad substrate specificity.

中文翻译：

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疏水性氨基酸转运蛋白 MhsT 跨膜螺旋 6 中的非螺旋区域介导底物识别


嗜盐芽孢杆菌的 MhsT 是疏水性氨基酸的转运蛋白，是氨基酸、神经递质、渗透剂或肌酸的 Na ⁺依赖性同向转运蛋白真核 SLC6 家族的同源物。 MhsT 转运的氨基酸范围广泛，促进了对底物识别机制的研究。在这里，我们报告了 MhsT 的六种新的底物结合结构，这些结构与功能研究相结合，揭示了跨膜片段 6 (TM6) 未卷绕区域中的 Gly-Met-Gly (GMG) 基序的灵活性如何对于通过定制结合位点的形状和体积来识别不同尺寸的底物。 MhsT 突变体在未缠绕的 GMG 环和底物结合口袋中包含取代，模仿中性氨基酸的真核 SLC6A18/B0AT3 和 SLC6A19/B0AT1 转运蛋白的结合位点，表现出需要大结合位点体积的芳香族氨基酸的转运受损。 SLC6 家族真核生物成员中一般 (G/A/C)ΦG 基序的保守性表明该环在具有广泛底物特异性的 SLC6 转运蛋白的底物识别和易位的常见机制中发挥作用。