Document Type

Thesis

Date of Award

5-2005

School/College

College of Science, Engineering, and Technology (COSET)

Degree Name

MS in Biology

First Advisor

Marian Hillar

Abstract

N, N-Dimethylformamide has been chosen as a potential effector to assess its allosteric influence on the enzymatic kinetics of the GDH enzyme. This study is part of a larger project designed toward determining the kinetic mechanism of glutamate dehydrogenase (GDH). The purpose of this study was to characterize the effects of N,N-Dimethylformamide (DMF) on the forward reaction of deamination of L-glutamic acid and build a mathematical model describing the enzymatic kinetic behavior. To determine these effects, reaction assays of NAD+ alone and in the presence of ADP and/or N, N-Dimethylformamide were recorded at 340 nm. All measurements were repeated several times (3-6) and the data displayed represented quantitative averages of all trials. Microsoft Excel software was used to graphically analyze experimental data. This study concluded that (1) the GDH enzyme displayed negative allosterism; (2) ADP abolishes the negative allosterism of GDH; (3) DMF inhibited GDH as a pure 1 2 competitive inhibitor; (4) Adair-Pauling fit provided the best curve-fitting model for GOH with the following allosteric interaction coefficients: a = 8.5; b = 1.8; c = 1.2; d = 2.3; and e = 7; and (5) OMF is an inhibitor of the enzyme reaction competing -for the low affinity NAO+ binding site (active site or Site I) specific for the amide moiety of-the-coenzyme

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