Date of Award

2012

Document Type

Thesis

Degree Name

Bachelors

Department

Natural Sciences

First Advisor

Walstrom, Katherine

Keywords

c. elegans, Kinetics, Biochemistry

Area of Concentration

Biochemistry

Abstract

Glyceraldehyde-3-phosphate dehydrogenase (GPD) is an enzyme which converts glyceraldehyde-3-phosphate (G3P) into 1,3-bisphosphoglycerate using NAD+ and Pi as substrates. Due to the involvement of GAPDH in glycolysis, it can be found almost universally among organisms. C. elegans contains 4 GPD genes, and these proteins have not been characterized kinetically. GPD-3 was chosen for this study due to its involvement in longevity in adult worms. The gpd-3 gene was inserted into the vector pTYB11, and the GPD-3 protein was overexpressed in E. coli cells. The GPD-3 purification was unsuccessful due to inefficient cleavage of the purification tag, so a preparation of endogenous GPD-2/GPD-3 was kinetically characterized. A Km of 0.3 mM and ~1 mM was obtained for NAD+ and G3P, respectively. These Km values are significantly higher than in other species studied, and suggest the potential for further analysis.

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