Substrate Specificity of a Methyltransferase Involved in the Biosynthesis of the Lantibiotic Cacaoidin

Modification of the N- and C-termini of peptides enhances their stability against degradation by exopeptidases. The biosynthetic pathways of many peptidic natural products feature enzymatic modification of their termini, and these enzymes may represent a valuable pool of biocatalysts. The lantibiotic cacaoidin carries an <i>N</i>,<i>N</i>-dimethylated N-terminal amine group. Its biosynthetic gene cluster encodes the putative methyltransferase Cao4. In this work, we present reconstitution of the activity of the enzyme, which we termed CaoS_C following standardized lanthipeptide nomenclature, using a heterologously produced peptide as the model substrate. In vitro methylation of diverse lanthipeptides revealed the substrate requirements of CaoS_C. The enzyme accepts peptides of varying lengths and C-terminal sequences but requires dehydroalanine or dehydrobutyrine at the second position. CaoS_C-mediated dimethylation of natural lantibiotics resulted in modestly enhanced antimicrobial activity of the lantibiotic haloduracin compared to that of the native compound. Improved activity and/or metabolic stability as a result of methylation illustrates the potential future application of CaoS_C in the bioengineering of therapeutic peptides.