Genome mining and genetic analysis of cypemycin biosynthesis reveal an unusual class of posttranslationally modified peptides

Abstract

Posttranslational modification of amino acids confers a range of structural features and activities on ribosomally synthesized peptides, many of which have potent antimicrobial or other biological activities. Cypemycin is an extensively modified linear peptide produced by Streptomyces sp. OH-4156 with potent in vitro activity against mouse leukemia cells. Cypemycin does not contain lanthionine bridges but exhibits some of the structural features of lantibiotics, notably dehydrated threonines (dehydrobutyrines) and a C-terminal S-[(Z)-2-aminovinyl]-D-cysteine. Consequently it was classified as a member of the lantibiotic family of posttranslationally modified peptides. Cypemycin also possesses two L-allo-isoleucine residues and an N-terminal N,N-dimethylalanine, both unique amino acid modifications. We identified and heterologously expressed the cypemycin biosynthetic gene cluster and performed a mutational analysis of each individual gene. We show that even the previously described modifications are carried out by unusual enzymes or via a modification pathway unrelated to lantibiotic biosynthesis. Bioinformatic analysis revealed the widespread occurrence of cypemycinlike gene clusters within the bacterial kingdom and in the Archaea. Cypemycin is the founding member of an unusual class of posttranslationally modified ribosomally synthesized peptides, the linaridins.Actinomycetes | antibiotic | enzymology | natural products | Streptomyces

Taxonomy, Physiology, and Natural Products of Actinobacteria

“…This is particularly important for heterologous expression of biosynthetic gene clusters that are being uncovered at a high rate in actinomycetes in the era of genome sequencing. In particular, the combination of synthetic biology approaches with expression in optimized heterologous Streptomyces production platforms is a promising development (403)(404)(405)(406)(407)(408).…”

Primer on Agar-Based Microbial Imaging Mass Spectrometry

“…Oftentimes, the distribution of the ion of interest is interfacial and the producer is ambiguous. By employing analytical and biological approaches, such as microbial IMS, IMS of multiple time points (67), traditional solvent extraction, purification, and structural determination methods, such as tandem MS and nuclear magnetic resonance (NMR) (18,30,33,67,68), genetics and microbiology, 16S rRNA sequencing (54), established MALDI-TOF protocols (12,49,53), genome mining approaches (7,13,15,45,63) and predictive programs (2,3,16,33, 40,48,55,62,65,68), peptidogenomics (31), and literature and database searches (23, 52) (AntiMarin database, Dictionary of Natural Products, the National Institute of Standards and Technology [NIST] databases, and the SciFinder database), one can typically annotate microbial IMS data. Two main strategies to confirm the producing microbe and the resultant phenotype are genetic knockout and complementation studies or assays with purified compound, in combination with more IMS.…”

Biosynthesis and Regulation of Grisemycin, a New Member of the Linaridin Family of Ribosomally Synthesized Peptides Produced by Streptomyces griseus IFO 13350

“…Several new classes of posttranslationally modified peptides have been identified based on recent advances in microbial genome sequencing, biochemistry, and genetics. Examples include the thiopeptides (9,15,17,26), patellamides (24), linear heterocyclized toxins (13), and linaridins (3). Interest in these compounds stems from the identification of previously undescribed peptides with potentially interesting biological properties and the discovery and characterization of new types of modification enzymes.…”