O-Methyltransferase-Mediated Incorporation of a β-Amino Acid in Lanthipeptides

Abstract

Lanthipeptides represent a large class of cyclic natural products defined by the presence of lanthionine (Lan) and methyllanthionine (MeLan) cross-links. With the advances in DNA sequencing technologies and genome mining tools, new biosynthetic enzymes capable of installing unusual structural features are continuously being discovered. In this study, we investigated an O-methyltransferase that is a member of the most prominent auxiliary enzyme family associated with class I lanthipeptide biosynthetic gene clusters. Despite the prevalence of these enzymes, their function has not been established. Herein, we demonstrate that the Omethyltransferase OlvS A encoded in the olv gene cluster from Streptomyces olivaceus NRRL B-3009 catalyzes the rearrangement of a highly conserved aspartate residue to a β-amino acid, isoaspartate, in the lanthipeptide OlvA(BCS A ). We elucidated the NMR solution structure of the GluC-digested peptide, OlvA(BCS A ) GluC , which revealed a unique ring topology comprising four interlocking rings and positions the isoaspartate residue in a solvent exposed loop that is stabilized by a MeLan ring. Gas chromatography−mass spectrometry analysis further indicated that OlvA(BCS A ) contains two DL-MeLan rings and two Lan rings with an unusual LL-stereochemistry. Lastly, in vitro reconstitution of OlvS A activity showed that it is a leader peptideindependent and S-adenosyl methionine-dependent O-methyltransferase that mediates the conversion of a highly conserved aspartate residue in a cyclic substrate into a succinimide, which is hydrolyzed to generate an Asp or isoAsp containing peptide. This overall transformation converts an α-amino acid into a β-amino acid in a ribosomally synthesized peptide, via an electrophilic intermediate that may be the intended product.

Precursor peptide-targeted mining of more than one hundred thousand genomes expands the lanthipeptide natural product family

“…The four largest class I families (I 1-4) are from Actinobacteria and do not have a characterized member. Their core peptides contain a highly conserved Asp residue that is of particular note because the corresponding BGCs contain an O-methyltransferase (PF01135) and the conserved Asp is likely post-translationally modified [46]. A number of the class II families, such as II 2, II 13, II 17, and II 26 have conserved leader peptides and non-conserved core peptides.…”

“…Methylation led to the formation of a succinimide that was hydrolyzed to a mixture of aspartate and isoaspartate. The mature form of the natural product remains unknown [46].…”

Precursor peptide-targeted mining of more than one hundred thousand genomes expands the lanthipeptide natural product family

“…The four largest class I families (I 1-4) are from Actinobacteria and do not have a characterized member. Their core peptides contain a highly conserved Asp residue that is of particular note because the corresponding BGCs contain an O-methyltransferase (PF01135) and the conserved Asp is likely posttranslationally modified [54]. A number of the class II families, such as II 2, II 13, II 17, and II 26 have conserved leader peptides and non-conserved core peptides.…”

Structure Prediction and Synthesis of a New Class of Macrocyclic Peptide Natural Products

“…Together, these advances have led to the discovery of numerous new natural products [24][25][26][27][28] and, in the case of RiPPs, new post-translational modifications. 14,[29][30][31][32][33][34][35] Bioinformatics recently has been utilized to discover new RiPP classes, such as the α-keto β-amino acid-containing peptides, 36 RiPPs generating thiaglutamate by peptide-amino acyl tRNA ligases (PEARLs), 33 aliphatic ether-containing rotapeptides 31 and non-α thioether-containing ranthipeptides, 37 as well as the pyritides, described herein. Bioinformatics offers a means to prioritize the characterization of divergent BGCs that give rise to novel compounds while also mitigating the historical issue of rediscovery.…”