Unveiling Marine Lanthipeptides: From Biosynthesis to Antibiotic Potential Against Antimicrobial Resistant Pathogens

PROJECT SUMMARY Nature remains the most lucrative resource for identifying new bioactive molecules for pharmacological applications. Identification of unique chemical scaffolds and new bacterial reservoirs for antibacterial natural products is required to reinvigorate the antibiotic discovery pipeline. Lanthipeptides are a class of natural products that frequently display potent antimicrobial activity with low susceptibility to antimicrobial resistance mechanisms, but an incomplete understanding of their biosynthesis and structure function relationships is creating a bottleneck between their antimicrobial potential and application as antibiotic therapies. Broader characterization of novel lanthipeptide scaffolds and the enzymes catalyzing their biosynthesis is needed to convert this untapped reservoir of bioactive natural products into antibiotic therapies. Genome mining approaches enable facile identification of an organism’s biosynthetic potential and have been used to demonstrate the importance of marine bacteria as untapped reservoirs for natural product discovery, including novel lanthipeptides. Two adjacent lanthipeptide biosynthetic gene clusters in an ecologically important species of marine bacteria were identified, with one of these clusters encoding for a predicted halogenase, which is a highly valuable catalyst in medical chemistry. The objective of this proposal is to structurally elucidate the lanthipeptide products produced by these novel biosynthetic gene clusters, test their antibiotic potential, and determine the substrate specificity of the halogenase involved in their biosynthesis. Indeed, the long-term goal is to contribute to the development of a molecular toolkit that can be used to engineer lanthipeptides into “unnatural natural products” with antibiotic activity. In Aim 1, ecological conditions resulting from host-microbial interactions will be used to trigger lanthipeptide production from the native marine bacterial species, allowing comparative metabolomics to be used to infer their chemical composition from wild type culture supernatants compared to deletion mutants. In Aim 2, the halogenase encoded within one of the lanthipeptide biosynthetic gene clusters will be co-expressed with wild type and mutated lanthipeptide precursor peptides along with their cognate biosynthesis enzymes. The resulting tailored peptides will be subject to chemical analysis to determine the range of substrate specificity of the tailoring enzyme. In Aim 3, lanthipeptide products, at varying stages of maturity, will be tested for antimicrobial activity against a panel of Gram positive and negative bacteria, and their mechanisms of action determined, as well as their suitability to act as adjuvants to restore antibiotic potency and reverse resistance. This proposal design embraces resources from a marine microbe whose well-established chemically mediated microbiome interactions have likely contributed to the evolution of new lanthipeptide scaffolds and halogenation machinery that can be exploited for new to nature pharmacophores.