Albonoursin

Two new piperazine-triones lansai E and F (1, 2), together with four known secondary metabolites lansai D (3), 1-N-methyl-(E,Z)-albonoursin (4), imidazo[4,5-e]-1,2,4-triazine (5), and streptonigrin (6) were isolated from a deep-sea-derived Streptomycetes sp. strain SMS636. The structures of the isolated compounds were confirmed by comprehensive spectroscopic analysis, including HRESIMS, 1D and 2D NMR. Compound 4 exhibited moderate antibacterial activities against Staphylococcus aureus and methicillin resistant S. aureus (MRSA) with Minimum Inhibitory Concentration (MIC) values of 12.5 and 25 μg/mL, respectively. Compound 6 displayed significant antibacterial activities against S. aureus, MRSA and Bacillus Calmette-Guérin (BCG) with MIC values of 0.78, 0.78 and 1.25 μg/mL, respectively.

Nocardiopsis alba is frequently isolated from environment and has recently been suggested as a casual symbiotic actinobacterium of diverse invertebrates. Using activity-guided fractionation, we purified two antibacterial cyclic dipeptides, cyclo(ΔPhe-ΔLeu) (albonoursin) and cyclo(ΔmTyr-ΔLeu), from a culture of Nocardiopsis alba ATCC BAA-2165. Analysis of N. alba genome revealed genetic information similar to albonoursin biosynthetic gene cluster, albABC. An albABC gene deletion mutant of N. alba was generated. Liquid chromatography-mass spectrometry analysis showed that the mutant could not produce the cyclic dipeptides. Cyclic dipeptide production in the mutant was restored by genetic complementation with the albABC cloned in a native plasmid of Nocardiopsis. β-Glucuronidase reporter assays with a second mutant construct, in which albABC promoter is transcriptionally fused to the reporting gene gusA, indicated that albABC gene expression was subject to osmoregulation. The system presented will be used to study the metabolic and genetic control of cyclic dipeptide biosynthesis in Nocardiopsis.

Cyclodipeptides and their derivatives belong to the diketopiperazine (DKP) family, which is comprised of a broad array of natural products that exhibit useful biological properties. In the few known DKP biosynthetic pathways, nonribosomal peptide synthetases (NRPSs) are involved in the synthesis of cyclodipeptides that constitute the DKP scaffold, except in the albonoursin (1) pathway. Albonoursin, or cyclo(alpha,beta-dehydroPhe-alpha,beta-dehydroLeu), is an antibacterial DKP produced by Streptomyces noursei. In this pathway, the formation of the cyclo(Phe-Leu) (2) intermediate is catalyzed by AlbC, a small protein unrelated to NRPSs. We demonstrated that AlbC uses aminoacyl-tRNAs as substrates to catalyze the formation of the DKP peptide bonds. Moreover, several other bacterial proteins, presenting moderate similarity to AlbC, also use aminoacyl-tRNAs to synthesize various cyclodipeptides. Therefore, AlbC and these related proteins belong to a newly defined family of enzymes that we have named cyclodipeptide synthases (CDPSs).