Abstract L-Threonine aldolase (L-TA) is a promising candidate for the optical synthesis of β-hydroxy-α-L-amino acids from glycine and kinds of aldehydes. Because of the Cβ-stereoselectivity problem, it was usually difficult to optically synthesize β-hydroxy-α-L-amino acids with Cβ-aromatic groups by application of L-TAs, especially those existing electron-donating groups on Cβ-aromatic groups, such as anti-Parkinson’s disease drug L-threo-DOPS (L-threo-3,4- dihydroxyphenylserine). A new L-TA (named Sz-1−2) from the gut microbiota of black bears was expressed and its Arg318 site-saturation mutation was conducted. Under the optimized conditions, three mutants R318 M, R318 V and R318 L showed 77.1 %, 79.9 % and 84.9 % de values respectively, which significantly improved than the wild-type (31.2 % de). Importantly, the mutant R318 L provided by far the best diastereoselectivity in the synthesis of L-threo-DOPS, 1.5 folds higher than the best-ever enzyme (55.4 % de). The L-TA R318 L is a valuable enzyme for the synthesis of L-threo-DOPS and might also be promising for the enzymatic synthesis of other important β-hydroxy-α-L-amino acids. Moreover, our research also suggested Arg318 was a crucial residue for the improvement of the diastereoselectivity of L-TAs, which could provide valuable experience for the relevant studies.

中文翻译：

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一种在合成 L-苏氨酸二羟基苯基丝氨酸中具有高非对映选择性的重组 L-苏氨酸醛缩酶

摘要 L-苏氨酸醛缩酶（L-TA）是一种从甘氨酸和多种醛类中光学合成β-羟基-α-L-氨基酸的有前途的候选者。由于Cβ-立体选择性问题，通常很难通过应用L-TAs来光学合成具有Cβ-芳基的β-羟基-α-L-氨基酸，尤其是那些存在于Cβ-芳基上的给电子基团，如抗帕金森病药物L-threo-DOPS（L-threo-3,4-dihydroxyphenylserine）。表达了一种来自黑熊肠道微生物群的新 L-TA（命名为 Sz-1-2），并进行了其 Arg318 位点饱和突变。在优化条件下，三个突变体R318 M、R318 V和R318 L分别表现出77.1 %、79.9 %和84.9 %的de值，显着高于野生型（31.2 % de）。重要的，迄今为止，突变体 R318 L 在 L-threo-DOPS 的合成中提供了最佳的非对映选择性，比有史以来最好的酶 (55.4 % de) 高 1.5 倍。L-TA R318 L 是合成 L-threo-DOPS 的一种有价值的酶，也有望用于其他重要 β-羟基-α-L-氨基酸的酶促合成。此外，我们的研究还表明Arg318是提高L-TAs非对映选择性的关键残基，可为相关研究提供宝贵经验。