Thesis (Ph.D.)--Virginia Commonwealth University, 2009. / Prepared for: Dept. of Chemistry. Title from title-page of electronic thesis. Bibliography: leaves 144-153.

Thesis (Ph.D.)--University of Delaware, 2006. / Principal faculty advisor: Brian J. Bahnson, Dept. of Chemistry and Biochemistry. Includes bibliographical references.

No description available.

Secondary N-nitrosamines: diphenylnitrosamine (DPhNA), dimethylnitrosamine (DMNA), diethylnitrosamine (DENA), dipropylnitrosamine (DPNA), dibutylnitrosamine (DBNA), diethanolnitrosamine (DEtNA), methylnitrosoglycine (MNGly), nitrosopyrrolidine (NPyr), nitrosomorpholine (NMor) and nitrosopiperidine (NPip) were synthesised and their interaction with acetylcholinesterase (AChE) was investigated. Analyses of kinetic results show that DMNA (Ki=34.78 μM); DENA (Ki=54.24 μM); DPNA(Ki=60.36 μM); DBNA(Ki=95.54 μM); DEtNA(Ki=43.68 μM)MNGly (Ki=30.18 μM); NPip (Ki=123 μM); NPyr (Ki=66.07 μM), NMor (Ki=73.93 μM) and DPhNA (Ki=20.32 μM) are competitive and reversible inhibitors of acetylcholinesterase, with respect to the substrate, acetylthiocholine chloride, ATChCl. With time they act as irreversible covalent inhibitors with dipropy1nitrosamine producing 72% inactivation after 60 minutes. Scatchard analyses of f1uorometric titrations, (Kd=0.75mM-4.09mM); gel chromatography (Kd=O. 80mM-4. 60mM) and equilibrium dia1ysis (Kd=O. 71mM- 4.21mM) for MNG1y, DMNA, DEtNA, DENA, DPNA, NPyr, DSNA, NMor and NPip show that these compounds have weaker affinity for the enzyme, as compared to the much tightly binding aromatic DPhNA, Kd values (0.65mM, 0.68mM and 0.68mM) for fluorometric experiments, gel chromatography and equilibrium dialysis respectively. In all cases, the number of binding sites of acetylcholinesterase averaged to four.

Parkinson's disease (PD) is a chronic, progressive disorder of the central nervous system that affects approximately 1.5 million Americans. One of the principal pathological features of PD is dopamine deficiency in the substantia nigra of the brain. A key enzyme that has been associated with the neurodegeneration seen in PD is monoamine oxidase-B (MAO-B). Several inhibitors of this enzyme have resulted in neuroprotection in the mouse model of PD. One such compound is 7-nitroindazole (1). This thesis describes the synthesis and MAO activity of several indazolyl and benzimidazolyl prodrugs that are designed to release an enzyme inhibitor in the affected brain area. These studies have provided information regarding the nucleophilic aromatic substitutions of the ambident nucleophiles under consideration. We have also discovered a compound that releases the enzyme inhibitor upon bioactivation by MAO. These results as well as a MPTP mouse study with the aforementioned compound are detailed within. / Master of Science

No description available.

Recombinant giardial arginine deiminase (rADI) was characterized. The enzyme was found to have a specific activity of 12 U (mg protein)-1under at pH 7.4 and 1 mM arginine. The maximum velocity was 14.75 U (mg protein-1) and the KM was 0.167 mM. The specific activity and maximum velocity values are significantly lower than the values reported previously for giardial rADI, while the KM value is quite similar. The optimum pH for the giardial rADI was 6-9, a broad range compared to other arginine deiminases. Recombinant ADI is very specific in its binding specificity, with canavanine (KI 2.4 mM) and ornithine (KI 2.1 mM) being the only substrate analogues giving significant inhibition from the wide variety of analogues tested. None of the analogues could be shown to act as alternative substrates. The contribution of conserved, catalytic and C-terminal residues proposed by previous research towards ADI activity was investigated by site-directed mutagenesis. Mutations of catalytic site residues Asp175, Glu226, His280 and Cys424 decreased the rADI activity to below 2%. Conservative mutations showed significant activity for Asp175 to Glu175 (DE175) and Glu226 to Asp226 (ED226). Site directed mutagenesis on the conserved non-catalytic site Leu46 showed activities below 15%, with the highest activity observed for the mutation to Val46 (LV46), which differs in one CH2 to Leu46 on its side chain. The KM of the mutant LV46 was 3.64 mM while for LA46 (Leu to Ala mutation) was 1.33 mM. Excising 5, 13, 16 amino acids from the C-terminal residues resulted in activity decreasing to 0.8% of the wild type, while excising 54 amino acids caused the rADI to be insoluble. Sequence alignment of Giardia and Dictyostelium suggests a homologous area within the C-terminal extension. Site directed mutagenesis on the Tyr567 residue in this region resulted in a decrease in activity, with the highest activity observed for a Tyr to Phe mutation. Studies using specific cysteine protease inhibitors demonstrated partial protection against proteolysis of ADI against giardial proteases. Degradation of ADI by giardial proteases was more rapid at pH 6 than at pH 7.4.

Graduation date: 1997

Human T-cell lymphotropic virus type I (HTLV-I) is a retrovirus that is the causative agent of the fatal disease adult T-cell leukemia (ATL). HTLV-I silently infects over twenty million people worldwide; up to ten percent of these will develop ATL in their lifetime. There are currently no effective treatments for this disease. HTLV-I expresses its genome as polypeptides that must be processed in order to produce infectious virions. Like other retroviruses, HTLV-I encodes an aspartic acid protease to process these polypeptides into mature form. Because the protease is essential in the virus life cycle, it is an attractive target for the treatment of HTLV-I-induced ATL. The present work examines the structure and function of HTLV-I protease. A theoretical structure of the protease is presented, and the function of the C-terminal extension is considered. In order to determine which residues are involved in binding substrate, two experiments were performed: first, several residues were mutated to the corresponding residues in HIV-1 protease to determine whether HTLV-I protease can be made to process an HIV-1 protease substrate; second, an alanine scan was performed to knock out individual residues to assess their importance in binding substrate. This work builds knowledge of the structure and function of HTLV-I protease. By understanding which residues play a role in binding substrate and by developing a clearer picture of the structure of the protease, it will be possible to develop specific inhibitors for HTLV-I protease.

Thesis (Ph. D.)--University of Washington, 2001. / Vita. Includes bibliographical references (leaves 76-85).