Competition between HPteGlu and HPtePAS Confers -Aminosalicylic Acid Resistance in .

Tuberculosis remains a serious challenge to human health worldwide. -Aminosalicylic acid (PAS) is an important anti-tuberculosis drug, which requires sequential activation by () dihydropteroate synthase and dihydrofolate synthase (DHFS, FolC). Previous studies showed that loss of function mutations of a thymidylate synthase coding gene caused PAS resistance in , but the mechanism is unclear. Here we showed that deleting in resulted in increased content of tetrahydrofolate (HPteGlu) in bacterial cells as they rely on the other thymidylate synthase ThyX to synthesize thymidylate, which produces HPteGlu during the process. Subsequently, data of in vitro enzymatic activity experiments showed that HPteGlu hinders PAS activation by competing with hydroxy dihydropteroate (HPtePAS) for FolC catalysis. Meanwhile, over-expressing in Δ strain and a PAS resistant clinical isolate with known mutation partially restored PAS sensitivity, which relieved the competition between HPteGlu and HPtePAS. Thus, loss of function mutations in led to increased HPteGlu content in bacterial cells, which competed with HPtePAS for catalysis by FolC and hence hindered the activation of PAS, leading to decreased production of hydroxyl dihydrofolate (HPtePAS-Glu) and finally caused PAS resistance. On the other hand, functional deficiency of in pushes the bacterium switch to an unidentified dihydrofolate reductase for HPteGlu biosynthesis, which might also contribute to the PAS resistance phenotype. Our study revealed how mutations confer PAS resistance in and provided new insights into studies on the folate metabolism of the bacterium.