SectionThe Corynebacterineae suborder encompasses many infectious agents, including deadly humanpathogens, such as Mycobacterium tuberculosis (M. tb). Many antibiotics used to treat M. tbblock steps in cell envelope assembly, underscoring the key role of this protective barrier. TheCorynebacterineae cell envelope is unique in that it is constructed from saccharide and lipidbuilding blocks that differ dramatically even from those used by other bacteria. One of thecentral features of the Corynebacterineae cell envelope is the mycolyl arabinogalactan (mAG),which is a glycoconjugate assembled from galactofuranose (Galf), arabinofuranose (Araf), andmycolic acids. Most enzymes involved in mAG biosynthesis have been identified but when,where, and how they function in live cells to construct and remodel the cell envelope is notknown. To elucidate these changes and their functional roles, the proposed aims focus ondeveloping new tools to probe, perturb, and exploit changes in the Corynebacterineae cellenvelope. The focus of Aim 1 is on generating a small molecule probe of the galactan?the onlymajor cell wall constituent for which no effective probes exist. The target enzyme is UDP-galactopyranose mutase (UGM), which catalyzes the formation of the key building blockrequired for biosynthesis of the essential galactan. The small molecule probe that results can beused to evaluate the consequences of blocking galactan assembly to understand the role ofgalactan in the stability and integrity of the cell envelope in Corynebacterineae and to probe therole of UGM in other organisms.
Aim 3 focuses on the mycolyltransferases, which construct thecritical mycolic acid components of the cell envelope. The localization of these enzymes and thetiming of their activity in cell division is not known. To visualize mycolyltransferase activity inlive cells, we propose to devise fluorogenic probes that can pinpoint the sites of Ag85 activityand reveal the interplay between mycolyltransferase activity and cell envelope assembly. Weanticipate that the probes described in Aim 3 can be used to develop assays for finding inhibitorsof cell division and for monitoring and detecting mycobacteria in real time. We expect that ourpursuit of the three aims shall uncover vulnerabilities in the defenses of mycobacteria andcorynebacteria that will lead to new antibiotic strategies.Significance:The overall objective of this application is to develop new chemical probes to understand howthe Corynebacterineae build and maintain their cell wall to survive. We anticipate that thisknowledge will lead ultimately to the identification of new strategies to treat infectious disease.
New strategies are needed to attack the growing prevalence of drug-resistant strains of bacteria;including Mycobacterium tuberculosis; the causative agent of tuberculosis. Because of theirunique cell wall; mycobacteria can survive under conditions that would be deadly for mostmicrobes; therefore; the focus of this project is on understanding how mycobacteria and relatedspecies build and remodel their cell wall. The identification of weak links in the cell wall can leadto antimycobacterial strategies that exploit these vulnerabilities.
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