Investigations of lanthipeptides derived from human gut microbiome

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Project Summary/Abstract The emergence of antibiotic resistance poses a serious global threat of growing concern to human, animal and environmental health. Therefore, there is an urgent need to discover alternative sources of new antibiotics to combat pathogenic infections while limiting the emergence of bacterial resistance. The van der Donk group focuses on the study of lanthipeptides, a class of ribosomally synthesized and posttranslationally modified peptide (RiPP) natural products characterized by lanthionine and methyllanthionine rings that have potential as novel pharmaceuticals and antimicrobial agents. Indeed, the lanthipeptide nisin has been utilized by the U.S. food industry as a preservative to combat food-borne pathogens for more than 50 years with little to no development of bacterial resistance. The human microbiome is a rich source of novel RiPP-like secondary metabolites with potent antimicrobial properties. However, the identities of nearly all these natural products are unknown, and their biological roles are poorly understood. Thus, a deeper analysis of human microbiome-derived natural products would not only lead to the discovery of new antimicrobial compounds but also guide new therapies that seek to modulate the human microbiota in order to impact human health. Through genome mining, three novel Clostridiales RiPP biosynthetic gene clusters (BGC) from Lachnospiraceae bacterium C6A11 (lah), Clostridium indicum (cli) and Clostridium sp. AF15-17LB (cls) were identified. Lachnospiraceae bacterium C6A11 colonizes the human gut and other Lachnospiraceae bacterium are linked to type 2 diabetes, and resistance to gut colonization by Clostridium difficile - a leading cause of nosocomial antibiotic-associated diarrhea. The BGCs are unique in that they encode an intriguing constellation of posttranslational modification enzymes (PTM) in conjunction with several distinct precursor peptides from which the natural products are derived. However, the identities of all these natural products are unknown and their biosynthesis poorly understood. Therefore, the research proposed herein is designed to examine all three BGCs by first developing a new generally applicable platform for accessing these natural products from E. coli and/or C. sporogenes for characterization and bioactivity screening. In addition, the role of each PTM enzyme will be examined through systematic gene deletions. The structure-function relationships of the PTM enzymes will be examined via protein crystallography and a subset will be bioengineered towards new functionalities. Completion of the proposed research will lead to the discovery of up to eleven novel RiPP natural products with potential therapeutic applications/interventions and the development of a new generally applicable platform for accessing other RiPP-like natural products from the human gut microbiome. In addition, the findings from this research will provide a platform of tools, knowledge and techniques that will extend to future discovery of new antibiotics and will form a blueprint for predicting the identity other similarly complicated BGCs from the human gut microbiome.