Mode of Action of Dithiolopyrrolone Antibiotics

Natural products made by bacteria have long been utilized as inspiration for antibiotic treatments. The growing threat of antimicrobial resistance has highlighted the critical need for new antibiotics which target bacteria with novel modes of action. The dithiolopyrrolone class of natural product antibiotics have been known since the mid 1900s. However, their modes of action and biosynthesis have remained elusive. We sought to better understand both the biosynthesis and mode of action of the dithiolopyrrolone antibiotics, holomycin and thiomarinol. Holomycin is produced by Streptomyces clavuligerus. The antibiotic is effective against both Gram-positive and Gram-negative bacteria and appears to have pleiotropic effects on bacteria. We find that holomycin in its reduced form is capable of both chelating metal ions and stripping zinc from enzyme active sites. We describe a mode of action for holomycin that unites these effects as originating from the disruption of metal homeostasis in Escherichia coli. Thiomarinol is another dithiolopyrrolone antibiotic whose mode of action is not fully understood. Thiomarinol is unique in that it is a hybrid antibiotic, composed of the pseudomonic acid analogue, marinolic acid, and the dithiolopyrrolone core, holothin. Bacterial isoleucyl tRNA synthetase is known as the target of pseudomonic acid and is also a likely target of thiomarinol as well. We show that both thiomarinol A and pseudomonic acid A inhibit isoleucine adenylation by Staphylococcus aureus isoleucyl tRNA synthetase with the same efficacy. We propose that either the dithiolopyrrolone portion of the antibiotic is promoting the uptake or retention of the compound in bacterial cells, or thiomarinol has additional inhibitory activities. Lastly, we create a heterologous production system for holomycin to study the antibiotic’s biosynthesis. Understanding the biosynthesis of holomycin can shed light on future chemoenzymatic derivatizations of the antibiotic with potentially different bioactivity.Doctor of Philosoph