Investigation of F420-dependent glucose-6-phosphate dehydrogenase variants using pH profiles and stopped-flow spectrometric ligand-binding methods

Author

Ana Laura Alvarez

ORCID Identifier(s)

0000-0002-9728-8113

Graduation Semester and Year

2022

Language

English

Document Type

Thesis

Degree Name

Master of Science in Chemistry

Department

Chemistry and Biochemistry

First Advisor

Kayunta Johnson-Winters

Abstract

F420-dependent glucose-6-phosphate dehydrogenase (FGD) from Mycobacteria tuberculosis, catalyzes a reaction the conversion of glucose-6-phosphate (G6P) to 6-phosphogluconolactone using the oxidized F420 cofactor, which is reduced to F420H2. The reaction is important within M. tuberculosis, the cause of tuberculosis disease (TB) due to its health relevance for treatment of multiple drug resistant and extreme drug resistant strains of M. tuberculosis. TB affects millions of people worldwide, and although treatable, multiple drug resistant and extreme drug resistant forms of TB adds strain on the current generation of drugs. Since this enzyme is not found in humans, is an ideal drug target. A past crystal structure of wild-type FGD from M. tuberculosis proposed that among the conserved residues, H40 and E109 function as the active site base and active site acid, respectively. In contrast, later pH profile studies on Glu109 and His 40 revealed that the while Glu109 acts as the acid, His 40 does not act as an active site base. Hence, our current focus is to determine which active site residue may act as active site base using the FGD variants of E13, H260 and H40. pH dependence studies were conducted to elucidate their roles in catalysis. The time-dependent binding experiments for wtFGD, E13, H40 were performed to get more mechanistic information such as koff and kon to incorporate into our global analyses. This work was conducted in collaboration with other group members and will be discussed here.

Keywords

F420-dependent enzymes, Kinetic analysis

Disciplines

Chemistry | Physical Sciences and Mathematics

License

This work is licensed under a Creative Commons Attribution-NonCommercial-Share Alike 4.0 International License.

Comments

Degree granted by The University of Texas at Arlington

Recommended Citation

Alvarez, Ana Laura, "Investigation of F420-dependent glucose-6-phosphate dehydrogenase variants using pH profiles and stopped-flow spectrometric ligand-binding methods" (2022). Chemistry & Biochemistry Theses. 85.
https://mavmatrix.uta.edu/chemistry_theses/85