Structural analysis of 1-deoxy-D-xylulose 5-phosphate synthase from Pseudomonas aeruginosa and Klebsiella pneumoniae reveals conformational changes upon cofactor binding

Isoprenoid precursor biosynthesis is an essential part of primary metabolism in all living organisms. While eukaryotes utilize the mevalonate (MEV) pathway for isoprenoid-precursor biosynthesis, the most important bacterial pathogens rely on the methylerythritol-phosphate (MEP) pathway. Therefore, enzymes involved in the MEP pathway are potentially valuable targets for the development of novel antibacterials to tackle the current antimicrobial resistance crisis. Within the MEP pathway, the enzyme 1-deoxy-D-xylulose-5-phosphate synthase (DXPS) represents a crucial, rate-limiting first step and a branch point in the biosynthesis of the vitamins B1 and B6. Herein, we present two novel, high-resolution DXPS crystal structures of the important ESKAPE pathogens Pseudomonas aeruginosa and Klebsiella pneumoniae in both the co-factor-bound and apo forms. We demonstrate that the absence of the cofactor ThDP leads to a disordered loop close to the active site and may be important for the design of potent DXPS inhibitors, albeit being different in both structures. In addition, we report the complex structure of paDXPS with a fluoropyruvate adduct, shedding more light on the structural basis of DXPS catalysis. Lastly, we have determined a complex structure of paDXPS with a thiamine analogue, opening up a route for structure-based drug design of this essential enzyme of the MEP pathway.