Investigating the Role of Residues E57 and E168 in M. Tuberculosis IGP Synthase Catalysis
Presentation Type
Poster
Faculty Advisor
Nina Goodey
Access Type
Event
Start Date
26-4-2024 12:45 PM
End Date
26-4-2024 1:44 PM
Description
Tuberculosis is a disease that consistently affects the lives of millions across the globe. It is caused by a pathogen called Mycobacterium tuberculosis, which relies on the function of the enzyme indole-3-glycerol phosphate synthase (MtIGPS). This enzyme catalyzes the conversion of 1-(o-carboxyphenylamino)-1-deoxyribulose-5-phosphate (CdRP), its substrate, into indole-3-glycerol phosphate (IGP), the product. Based on previously determined solvent deuterium kinetic isotope effect data, our hypothesis is that the rate-limiting step in the enzyme-catalyzed reaction is a chemistry step that involves proton transfer. We also hypothesize that mutations to the enzyme (E57N, E57D, E168D) may result in changes to this step. For example, it is possible that some of these mutants have a rate-limiting substrate binding or product release step. Moreover, we suspect that the rate-pH profile of MtIGPS will change as a result of the mutations because residue E57 forms a hydrogen bond with the putative catalytic acid Lys119, and E168 may serve as a catalytic base in the mechanism. The MtIGPS mutants E57N, E57D, and E168D are expressed and their purity and concentrations are analyzed. Next we determine their rate vs pH profiles to ascertain the optimal pH for catalytic activity and the shape of the rate-pH curve for each of the mutants. As tuberculosis continues to show increasing resistance against a multitude of drugs, it is possible that new mechanism based inhibitors can be designed to specifically target the MtIGPS enzyme.
Investigating the Role of Residues E57 and E168 in M. Tuberculosis IGP Synthase Catalysis
Tuberculosis is a disease that consistently affects the lives of millions across the globe. It is caused by a pathogen called Mycobacterium tuberculosis, which relies on the function of the enzyme indole-3-glycerol phosphate synthase (MtIGPS). This enzyme catalyzes the conversion of 1-(o-carboxyphenylamino)-1-deoxyribulose-5-phosphate (CdRP), its substrate, into indole-3-glycerol phosphate (IGP), the product. Based on previously determined solvent deuterium kinetic isotope effect data, our hypothesis is that the rate-limiting step in the enzyme-catalyzed reaction is a chemistry step that involves proton transfer. We also hypothesize that mutations to the enzyme (E57N, E57D, E168D) may result in changes to this step. For example, it is possible that some of these mutants have a rate-limiting substrate binding or product release step. Moreover, we suspect that the rate-pH profile of MtIGPS will change as a result of the mutations because residue E57 forms a hydrogen bond with the putative catalytic acid Lys119, and E168 may serve as a catalytic base in the mechanism. The MtIGPS mutants E57N, E57D, and E168D are expressed and their purity and concentrations are analyzed. Next we determine their rate vs pH profiles to ascertain the optimal pH for catalytic activity and the shape of the rate-pH curve for each of the mutants. As tuberculosis continues to show increasing resistance against a multitude of drugs, it is possible that new mechanism based inhibitors can be designed to specifically target the MtIGPS enzyme.