Thiazolyl peptides possess novel scaffolds and proven activity against dangerous drug resistant pathogens such as methicillin-resistant Staphylococcus auerus (MRSA) and Plasmodium falciparum that make them ideal candidates for antibiotic development. Despite their potency and the pressing need for new antibiotics to combat emerging resistance, thiazolyl peptides have eluded development as therapeutics due to insufficient methods for their diversification. This deficiency has also hindered the study of structure-activity relationships to define how their novel scaffolds direct the antibiotic mode of action. The recent discovery that these molecules arise from ribosomal precursor peptides which undergo extensive post-translational modifications opens the door to rapid diversification through genetic manipulation. Towards this end, this research aims to create a novel thiazolyl peptide expression platform to enhance the understanding of how thiazolyl peptide scaffolds provide exquisite targeting of ribosomal protein synthesis in bacteria. Our specific hypothesis is that modified thiazolyl peptide ring structures will have alternate or multiple targets that inhibit the growth of gram positive bacteria. Preliminary data indicates the thiazolyl peptide biosynthetic machinery should readily accept modest changes in ring size. We intend to accomplish these goals through the following specific aims: 1. Identify the gene cluster responsible for the thiazolyl peptide GE37468 production in Streptomyces sp. ATCC 55365 and transfer it to the genetically tractable Streptomyces lividans to create a stable thiazolyl peptide expression platform. This will establish the first gene cluster from a publically accessible organism which produces a thiazolyl peptide with a 29-member ring and give insights into the biosynthesis of the unique GE37468 structure. 2. Create rational and high-diversity libraries of GE37468 analogs with altered ring sizes using the novel S. lividans expression platform. 3. Evaluate the mode of antibiotic activity for GE37468 analogs in a high through-put manner to create a better understanding of how structure provides targeting. The research proposed herein will take an unprecedented leap in the ability to create, understand, and screen libraries of cyclic peptides. This work has the potential to create thiazolyl peptides with novel spectrums of activity through the construction of high diversity genetic libraries. This is vital to the long-term goal of creating thiazolyl peptide therapeutics that can combat emerging resistance in pathogenic bacteria.
This project is relevant to public health because it proposes to develop greater understanding of how thiazolyl peptides inhibit protein synthesis in pathogenic bacteria. This understanding is vital to the development of new therapeutics to combat emerging bacterial resistance.
|Malcolmson, Steven J; Young, Travis S; Ruby, J Graham et al. (2013) The posttranslational modification cascade to the thiopeptide berninamycin generates linear forms and altered macrocyclic scaffolds. Proc Natl Acad Sci U S A 110:8483-8|