Thesis (Ph. D.)--University of Hong Kong, 1996. / Includes bibliographical references (leaf 185-194).

Thesis--Auburn University, 2009. / Abstract. Vita. Includes bibliographic references (p.49-58).

Thesis (Ph.D.)--University of Hong Kong, 1996. / Includes bibliographical references (leaf 185-194). Also available in print.

Würzburg, Univ., Diss., 2008. / Zsfassung in dt. Sprache.

Universiẗat, Diss., 2007--Frankfurt (Main). / Zsfassung in dt. und engl. Sprache.

No description available.

Malaria affects the daily lives of more than 2 billion people worldwide and has been estimated to result in 300-500 million clinical cases annually leading to approximately 2 million deaths, mainly caused by the most virulent malaria species, Plasmodium falciparum. The lack of a vaccine and the rapid emergence and spread of drug resistant strains of P. falciparum, necessitate the development of new antimalarials and the identification and validation of new parasite-specific therapeutic targets. Numerous studies directed at interfering with the polyamine biosynthetic pathway in P. falciparum have shown its potential as a target for the development of a new class of antimalarials. The essential nature of P. falciparum spermidine synthase (PfSpdSyn), an enzyme in the polyamine pathway of the parasite warranted the further investigation to find novel lead compounds. The high cost and attrition rate of drug discovery has resulted in the implementation of smart drug discovery platforms in both academia and industry. The strategy implemented in this study involved the development of a dynamic receptor-based pharmacophore model (DPM) of PfSpdSyn complemented by a knowledge-based rational design strategy. The use of pharmacophore models to identify lead compounds has become increasingly popular over the last decade and has been shown to be a reliable method in the drug discovery process. The development of a DPM allows for the incorporation of protein exibility within the drug design process. This methodology results in a wealth of information of the chemical space of the active site and was incorporated in designing new inhibitors against PfSpdSyn using a knowledge-based rational design strategy. The active site of PfSpdSyn was subdivided into four binding regions (DPM1-DPM4) to allow for the identi cation of fragments binding within these speci c binding regions. DPMs representative of the chemical characteristics of each binding region were constructed and subsequently screened against the drug-like subset of the ZINC database. From the screens a total of nine compounds were selected for in vitro testing, complementing each other in exploring specific active site binding characteristics. From these compounds a new lead compound N-(3-aminopropyl)-cyclohexylamine (NAC; Ki 2.8 μM) was identified for PfSpdSyn. NAC was specifically designed to bind in both the putrescine and decarboxylated adenosylmethionine cavities by chemically bridging the catalytic center and was confirmed by kinetic studies. NAC shows great potential for lead optimization to increase its binding affinity. This study then paves the way for lead optimization and possibly the development of a novel antimalarial. The development of a DPM for PfSpdSyn has seen the establishment of this methodology in the Bioinformatics and Computational Biology Unit, Department of Biochemistry at the University of Pretoria. It can be concluded that the development of a DPM complemented by a knowledge-based rational design strategy is an effective approach for the identification of novel lead compounds in the presence of a 3D target structure. This paves the way for more studies on both malaria and other drug targets using DPMs. Copyright / Thesis (PhD)--University of Pretoria, 2009. / Biochemistry / unrestricted

Hydroxymethylglutaryl-CoA synthase (HMG-CoA synthase) catalyzes the formation of HMG-CoA from acetyl-CoA and acetoacetyl-CoA. F-244 (1), a naturally occurring J3-lactone isolated from Fusarium sp. ATCC 20788 and other species, is known to be a potent and specific inhibitor ofHMG-CoA synthase isolated from rat liver. 1Ji,, HO,.,._.. .. , '• .. ,....13_.,., ;=( 0 14 0 0 (1) (2) This thesis describes the 48 fold purification of HMG-CoA synthase from bakers yeast in a three step procedure involving ethanol fractionation followed by ammonium sulfate precipitation and then hydroxylapatite chromatography. This procedure was found to be reproducible and yields a preparation of specific activity 0.14 units (j...tmolfmin)/mg in an overall yield of 8%. In our study, F-244 was found to be a potent irreversible inhibitor of HMGCoA synthase isolated from bakers yeast, with an IC50 value of 0.009 j...tM. This value is almost identical to the inhibitory activity of F-244 on rat liver HMG-CoA synthase that has been reported in the literature. Tritium labeled F-244 was prepared, for the first time, by feeding methyl-[3H]methionine to cultures of Fusarium sp. The [15, 16, 17, 18-3H] F-244 isolated had a specific activity of 1.3 x 106 DPM/mg. This tritiated F-244 was then used as an affinity.label for HMG-CoA synthase. Attempts to isolate the enzyme-inhibitor complex were unsuccessful due to the low level of radioactivity associated with the tritiated F-244. HMG-CoA synthase was also shown to be inhibited in a time-dependent irreversible manner by {±)-(3-butyrolactone (2). The rate of inactivation (~) was found to be 0.4697 s-1 and the inhibition constant (K1) was found to be 9 mM. The inactivation was found to be irreversible over several hours. / Thesis / Master of Science (MS)

In this paper we present the inhibitory effect of a variety of structurally modulated/modified polyphenolic compounds on purified F 1 or membrane bound F 1F o Escherichia coli ATP synthase. Structural modulation of polyphenols with two phenolic rings inhibited ATP synthase essentially completely; one or three ringed polyphenols individually or fused together inhibited partially. We found that the position of hydroxyl and nitro groups plays critical role in the degree of binding and inhibition of ATPase activity. The extended positioning of hydroxyl groups on imino diphenolic compounds diminished the inhibition and abridged position enhanced the inhibition potency. This was contrary to the effect by simple single ringed phenolic compounds where extended positioning of hydroxyl group was found to be effective for inhibition. Also, introduction of nitro group augmented the inhibition on molar scale in comparison to the inhibition by resveratrol but addition of phosphate group did not. Similarly, aromatic diol or triol with rigid or planar ring structure and no free rotation poorly inhibited the ATPase activity. The inhibition was identical in both F 1F o membrane preparations as well as in isolated purified F 1 and was reversible in all cases. Growth assays suggested that modulated compounds used in this study inhibited F 1-ATPase as well as ATP synthesis nearly equally.

published_or_final_version / Botany / Doctoral / Doctor of Philosophy