Recombinant Production of E. coli NAD+-dependent DNA ligase as a Target for Antibacterial Drug Discovery

The increase in the frequency of drug resistance in bacterial infections has led to the research of new antibacterial agents targeting new mechanisms. Many of the functions of NAD+-dependent DNA ligase have made it a remarkable target for antibacterial drug discovery. Escherichia coli (E. coli) NAD+-dependent DNA ligase is presented as a potential target due to its unique substrate specificity compared to the ATP-dependent human DNA ligase. In this study, it was aimed to produce and purify the E. coli NAD + dependent DNA ligase enzyme, which is frequently used in antibacterial drug discovery. The E. coli DNA ligase gene sequence was cloned into pTOLT vector system. E. coli DNA ligase enzyme was purified after the production in E. coli BL21 (DE3) pLysE cells. It was clearly demonstrated by the activity test that the DNA ligase enzyme produced in this study can ligate the DNA fragments. As a result, it was revealed that the effect of candidate inhibitors can be studied simply on the enzyme.