Tuberculosis (TB) is one of the world's deadliest diseases. Currently a third of the world's population is infected by Mycobacterium tuberculosis (MyTB), the causative agent for TB. In 2012 alone almost 9 million people developed TB resulting in 1.3 million deaths. Current antibiotic therapies have limited efficacy and must typically be used in combination and for prolonged periods of time. Furthermore, drug-resistant forms of MyTB are increasingly prevalent, underscoring the urgency to identify new targets and develop novel drugs against TB. The key determinants of all MyTB-host interactions are glycolipids in the cell wall of the pathogen that all share a common phosphatidylinositol (PI) anchor to the membrane and indeed PI is essential for MyTB survival. The crucial first step in biosynthesis of mycobacterial PI is carried out by an enzyme called phosphatidylinositolphosphate synthase (PIPS). PIPS belongs to a large family of enzymes called the CDP-alcohol phosphotransferases (CDP-AP). CDP-APs play the key role in the biosynthesis of all glycerophospholipids across all kingdoms of life. We have determined the atomic resolution structure by X-ray crystallography of the first representative CDP-AP and we now have solved a second structure of a PIPS which is 40% identical to the MyTB enzyme. Here we propose: (i) to use this PIPS model to study in molecular detail PI synthesis in MyTB (Aim 1); (ii) to build upon our expertise with CDP-APs to determine the structure of MyTB PIPS (Aim 2); (iii) to capitalize on our capability to generate abundant quantities of functional recombinant MyTB PIPS to measure its activity and set up a high-throughput screening assay for MyTB PIPS inhibitors whilst in parallel exploring chemical space by virtual screening to identify potential leads. Our research will not only offer unprecedented insight into how PI is made in MyTB, but it will also allow us to initiate structure-guided drug design efforts targeting the synthesis of this essential lipid. PIPS represents a key target for development of narrow spectrum anti- mycobacterial therapeutics. Indeed, the substrate for MyTB PIPS is unique to Archaea and few bacteria including Mycobacteria. In contrast, eukaryotic PI-synthases do not process inositol phosphate and most bacteria lack phosphatidylinositol entirely. We are in a unique position to set this framework given our results and expertise on the structures of CDP-APs and PIPS.
Tuberculosis (TB) causes well over 1 million deaths every year. We will contribute atomic resolution structures to develop drugs targeting phosphatidylinositol-phosphate synthase (PIPS), an essential enzyme in Mycobacterium tuberculosis, the causative agent for TB. Targeting this pathway is particularly appealing as Mycobacteria cannot survive without phosphatidylinositol (PI), most other bacteria don't have PI, eukaryotic PI biosynthesis proceeds through a different route, and we present as preliminary data the crystal structure of a PIPS to serve as a model for structure-guided drug design.
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