Aspirochlorine is an unusual antifungal cyclopeptide produced by Aspergillus oryzae,a ni mportant mold used for food fermentation. Whereas its structure suggested that anon-ribosomal peptide synthetase assembles the cyclopeptide from phenylalanine and glycine building blocks,l abeling studies indicated that one Phe moiety is transformed into Gly after peptide formation. By means of genetic engineering, heterologous expression, biotransformations,a nd in vitro assays,w ed issected and reconstituted four crucial steps in aspirochlorine biosynthesis,w hichi nvolve two cytochrome P450 monooxygenases,(AclL and AclO), amethyltransferase (AclU), and ah alogenase (AclH). We found that the installation of the N-methoxylation of the peptide bond sets the stage for ar etro-aldol reaction that leads to the Phe-to-Gly conversion. The substrate scopes of the dedicated enzymes as well as bioassays revealed that the peptide editing has evolved to optimize the antifungal action of the natural product.
Our observation suggests that detection of active MMP-2 in pancreatic juice using gelatin zymography may be useful for the diagnosis of pancreatic cancer.
To identify natural products and their associated biosynthetic genes from underutilized, difficult-to-manipulate microbes, chemical screening and bioinformatic analysis were employed to identify secondary metabolites and a potentially associated biosynthetic gene cluster. Subsequently, a heterologous expression system was used to confirm the identity of the gene cluster and the proposed biosynthetic mechanism. This approach successfully identified the curvupallide and spirostaphylotrichin biosynthetic pathways in endophytic fungus Curvularia pallescens and the short-chain pyranonigrin biosynthetic pathway in Aspergillus niger.
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