Bacterial MurF Assays
MurF is the enzyme that catalyzes the last step in synthesis of UDP-MurNAc-pentapeptide in the pathway of peptidoglycan biosynthesis in bacteria. It is an essential enzyme and attractive target for anti-bacterial drug discovery. MurF adds a dipeptide D-Ala-D-Ala onto the MurE product UDP-MurNAc-tripeptide. The ligation reaction is coupled to the hydrolysis of ATP forming ADP and inorganic phosphate.
The Bacterial MurF Assay is based on measurement of the inorganic phosphate generated from the MurF reaction. The inorganic phosphate is detected by light absorbance at 650 nm. The assay reactions and detection can be performed by using 384-well or 96-well assay plates. Alternatively, the assay reaction can be carried out in Eppendorf tubes and the signal is measured using a cuvette. The high throughput assay can be used for screening inhibitors of bacterial MurF in drug discovery research. It may also be used for characterization of bacterial MurF.