Pyrroloiminoquinone-containing natural products have long been known for their biological activities. They are derived from tryptophan, but their biosynthetic pathways have remained elusive. Studies on the biosynthetic gene cluster (BGC) that produces the ammosamides revealed that the first step is attachment of Trp to the C-terminus of a scaffold peptide in an ATP- and tRNA-dependent manner catalyzed by a PEptide Aminoacyl-tRNA Ligase (PEARL). The indole of Trp is then oxidized to a hydroxyquinone. We previously proposed a chemically plausible and streamlined pathway for converting this intermediate to the ammosamides using additional enzymes encoded in the BGC. In this study, we report the activity of four additional enzymes from two gene clusters, which show that the previously proposed pathway is incorrect and that Nature’s route toward pyrroloiminoquinones is much more complicated. We demonstrate that, surprisingly, amino groups in pyrroloiminoquinones are derived from (at least) three different sources, glycine, asparagine, and leucine, all introduced in a tRNA-dependent manner. We also show that an FAD-dependent putative glycine oxidase (Amm14) is required for the process that incorporates the nitrogens from glycine and leucine and that a quinone reductase is required for the incorporation of asparagine. Additionally, we provide the first insights into the evolutionary origin of the PEARLs as well as related enzymes, such as the glutamyl-tRNA-dependent dehydratases involved in the biosynthesis of lanthipeptides and thiopeptides. These enzymes appear to all have descended from the ATP-GRASP protein family.

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吡咯亚氨基醌生物合成过程中的意外转化

含吡咯亚氨基醌的天然产物长期以来因其生物活性而闻名。它们源自色氨酸，但其生物合成途径仍然难以捉摸。对产生氨酰胺的生物合成基因簇 (BGC) 的研究表明，第一步是在 PEeptide 氨酰基-tRNA 连接酶 (PEARL) 的催化下，以 ATP 和 tRNA 依赖性方式将 Trp 连接到支架肽的 C 末端。 ）。然后色氨酸的吲哚被氧化成羟基醌。我们之前提出了一种化学上合理且简化的途径，使用 BGC 中编码的其他酶将该中间体转化为氨酰胺。在这项研究中，我们报告了来自两个基因簇的另外四种酶的活性，这表明先前提出的途径是不正确的，并且自然界通往吡咯亚氨基醌的途径要复杂得多。令人惊讶的是，我们证明吡咯亚氨基醌中的氨基来自（至少）三种不同的来源：甘氨酸、天冬酰胺和亮氨酸，它们都是以 tRNA 依赖性方式引入的。我们还表明，结合甘氨酸和亮氨酸中的氮的过程需要依赖于 FAD 的推定甘氨酸氧化酶 (Amm14)，并且结合天冬酰胺需要醌还原酶。此外，我们首次深入了解 PEARL 以及相关酶的进化起源，例如参与羊毛硫肽和硫肽生物合成的谷氨酰-tRNA 依赖性脱水酶。这些酶似乎都源自 ATP-GRASP 蛋白家族。