Investigating the impact of viscosity on Mycobacterium Tuberculosis IGP synthase catalysis
Presentation Type
Abstract
Faculty Advisor
Nina Goodey
Access Type
Event
Start Date
25-4-2025 9:00 AM
End Date
25-4-2025 9:59 AM
Description
Tuberculosis affects the lives of millions throughout the world. Despite the efforts to treat tuberculosis with antibiotics, it is endlessly producing new strains that render drugs less effective. Indole-3-glycerol phosphate synthase (IGP synthase) catalyzes the conversion of 1-(o-carboxyphenylamino)-1-deoxyribulose-5-phosphate (CdRP) into indole-3-glycerol phosphate (IGP), which is the fourth step of the tryptophan biosynthetic pathway. Tryptophan is needed for the survival of Mycobacterium tuberculosis, the tuberculosis-causing pathogen. We aim to understand the IGP synthase catalyzed reaction with the goal of creating mechanism-based inhibitors. Such inhibitors will target the IGP synthase enzyme in Mycobacterium tuberculosis, stopping the production of IGP and tryptophan. For this purpose, the wildtype IGP Synthase and single-point mutant E168D were utilized in solvent viscosity effect (SVE) experiments. The concentrations of three viscogens (glycerol, sucrose, and polyethylene glycol 8000) were varied to investigate the effects on catalytic activity and substrate binding for the wildtype and E168D mutant. The results revealed the presence of an inverse SVE in E168D, with the enzymatic activity increasing from 0.0001 sec-1 to 0.0004 sec-1 as glycerol concentration increased. The activity doubled when in sucrose and polyethylene glycol, increasing from 0.0001 sec-1 to 0.0002 sec-1. In all three cases, the KM decreased, showing an increased affinity for the substrate as viscosity increased. This suggests the existence of a conformational change that is rate-limiting, following the binding of substrate to the enzyme. In knowing this, we may gain insight into the mechanism of IGP synthase to manufacture a novel inhibitor.
Investigating the impact of viscosity on Mycobacterium Tuberculosis IGP synthase catalysis
Tuberculosis affects the lives of millions throughout the world. Despite the efforts to treat tuberculosis with antibiotics, it is endlessly producing new strains that render drugs less effective. Indole-3-glycerol phosphate synthase (IGP synthase) catalyzes the conversion of 1-(o-carboxyphenylamino)-1-deoxyribulose-5-phosphate (CdRP) into indole-3-glycerol phosphate (IGP), which is the fourth step of the tryptophan biosynthetic pathway. Tryptophan is needed for the survival of Mycobacterium tuberculosis, the tuberculosis-causing pathogen. We aim to understand the IGP synthase catalyzed reaction with the goal of creating mechanism-based inhibitors. Such inhibitors will target the IGP synthase enzyme in Mycobacterium tuberculosis, stopping the production of IGP and tryptophan. For this purpose, the wildtype IGP Synthase and single-point mutant E168D were utilized in solvent viscosity effect (SVE) experiments. The concentrations of three viscogens (glycerol, sucrose, and polyethylene glycol 8000) were varied to investigate the effects on catalytic activity and substrate binding for the wildtype and E168D mutant. The results revealed the presence of an inverse SVE in E168D, with the enzymatic activity increasing from 0.0001 sec-1 to 0.0004 sec-1 as glycerol concentration increased. The activity doubled when in sucrose and polyethylene glycol, increasing from 0.0001 sec-1 to 0.0002 sec-1. In all three cases, the KM decreased, showing an increased affinity for the substrate as viscosity increased. This suggests the existence of a conformational change that is rate-limiting, following the binding of substrate to the enzyme. In knowing this, we may gain insight into the mechanism of IGP synthase to manufacture a novel inhibitor.
Comments
Poster presentation at the 2025 Student Research Symposium.