This study presents the crystal structure of a thiol variant of the human mitochondrial branched-chain aminotransferase protein. Human branched-chain aminotransferase (hBCAT) catalyzes the transamination of the branched-chain amino acids leucine, valine and isoleucine and α-ketoglutarate to their respective α-keto acids and glutamate. hBCAT activity is regulated by a CXXC center located approximately 10 Å from the active site. This redox-active center facilitates recycling between the reduced and oxidized states, representing hBCAT in its active and inactive forms, respectively. Site-directed mutagenesis of the redox sensor (Cys315) results in a significant loss of activity, with no loss of activity reported on the mutation of the resolving cysteine (Cys318), which allows the reversible formation of a disulfide bond between Cys315 and Cys318. The crystal structure of the oxidized form of the C318A variant was used to better understand the contributions of the individual cysteines and their oxidation states. The structure reveals the modified CXXC center in a conformation similar to that in the oxidized wild type, supporting the notion that its regulatory mechanism depends on switching the Cys315 side chain between active and inactive conformations. Moreover, the structure reveals conformational differences in the N-terminal and inter-domain region that may correlate with the inactivated state of the CXXC center.

中文翻译：

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人线粒体支链转氨酶氧化突变体的晶体结构。

这项研究展示了人线粒体支链转氨酶蛋白的硫醇变体的晶体结构。人支链转氨酶 (hBCAT) 催化支链氨基酸亮氨酸、缬氨酸和异亮氨酸以及 α-酮戊二酸转氨为各自的 α-酮酸和谷氨酸。hBCAT 活性由距离活性位点约 10 Å 的 CXXC 中心调节。该氧化还原活性中心促进还原态和氧化态之间的循环，分别代表活性和非活性形式的 hBCAT。氧化还原传感器 (Cys315) 的定点诱变导致活性显着丧失，但解析半胱氨酸 (Cys318) 的突变没有报告活性丧失，这允许在 Cys315 和 Cys318 之间可逆形成二硫键。C318A 变体氧化形式的晶体结构用于更好地了解各个半胱氨酸及其氧化态的贡献。该结构揭示了修饰后的 CXXC 中心的构象与氧化野生型相似，支持了其调节机制取决于 Cys315 侧链在活性和非活性构象之间切换的观点。此外，该结构揭示了 N 端和域间区域的构象差异，这可能与 CXXC 中心的失活状态相关。