Lincosamides are clinically important antibiotics originally produced as microbial specialized metabolites. The complex biosynthesis of lincosamides is coupled to the metabolism of mycothiol as a sulfur donor. Here, we elucidated the N-deacetylation of the mycothiol-derived N-acetyl-l-cysteine residue of a lincosamide intermediate, which is comprised of an amino acid and an aminooctose connected via an amide bond. We purified this intermediate from the culture broth of a deletion mutant strain and tested it as a substrate of recombinant lincosamide biosynthetic proteins in the in vitro assays that were monitored via liquid chromatography–mass spectrometry. Our findings showed that the N-deacetylation reaction is catalyzed by CcbIH/CcbQ or LmbIH/LmbQ proteins in celesticetin and lincomycin biosynthesis, respectively. These are the first N-deacetylases from the TldD/PmbA protein family, from which otherwise only several proteases and peptidases were functionally characterized. Furthermore, we present a sequence similarity network of TldD/PmbA proteins, which suggests that the lincosamide N-deacetylases are unique among these widely distributed proteins.

中文翻译：

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TcosD / PmbA家族蛋白催化Lincosamide生物合成中的N-脱乙酰化。

林可酰胺是临床上重要的抗生素，最初是作为微生物的专门代谢产物产生的。林可酰胺的复杂生物合成与作为硫供体的霉菌硫醇的代谢有关。在这里，我们阐明了Ñ的分支硫醇衍生-deacetylation Ñ乙酰基升-半胱氨酸林可酰胺中间体，其包含的氨基酸和通过酰胺键连接的aminooctose的残基。我们从缺失突变株的培养液中纯化了该中间体，并在通过液相色谱-质谱法监测的体外分析中将其作为重组林可酰胺生物合成蛋白的底物进行了测试。我们的研究结果表明，ñcelesticetin和林可霉素生物合成中的CcbIH / CcbQ或LmbIH / LmbQ蛋白分别催化脱乙酰反应。这些是来自TldD / PmbA蛋白家族的第一个N-脱乙酰基酶，否则仅从功能上鉴定了几种蛋白酶和肽酶。此外，我们提出了TldD / PmbA蛋白的序列相似性网络，这表明在这些广泛分布的蛋白中，林可酰胺N-脱乙酰酶是独特的。