SITE-DIRECTED DRUG DESIGN AGAINST PLASMODIUM FALCIPARUM.

THE ULTIMATE OBJECTIVE OF THIS PROJECT IS TO UTILIZE THE INFORMATION AVAILABLE IN THE THREE-DIMENSIONAL CRYSTAL STRUCTURE OF AN ENZYME IN THE RATIONAL DESIGN OF DRUGS WHICHWILL BE EFFECTIVE AGAINST PLASMODIUM FALCIPARUM. THE SPECIFIC ENZYME TARGET IS THE BIFUNCTIONAL DIHYDROFOLATE REDUCTASE-THYMIDYLATE SYNTHASE (DHFR-TS) OF P. FALCIPARUM. THE ACHIEVEMENT OF THIS GOAL WILL INVOLVE A CONCERTED EFFORTALONG SEVERAL FRONTS. FIRST, THE DHFR-TS GENE MUST BE ISOLATED AND CLONED INTO AN E. COLI EXPRESSION VECTOR WHICH WILL ALLOW ADEQUATE OVERPRODUCTION OF THE PROTEIN. SECOND, CHROMATOGRAPHIC PROCEDURES WILL BE DEVISED FOR THE ISOLATIONOF HOMOGENEOUS PROTEIN. THIRD, THE PROTEIN WILL BE CRYSTALLIZED AND ITS THREE-DIMENSIONAL STRUCTURE DISCERNED. FOURTH, THE THREE-DIMENSIONAL SPACE AT THE ACTIVE SITE WILL BE USED FOR THE DESIGN OF CHEMICAL COMPOUNDS WHICH WOULD BE EXPECTED TO BIND TIGHTLY AND BE STRONGLY INHIBITORY FOR THE P. FALCIPARUM DHFR-TS WHILE HAVING MUCH REDUCED AFFINITY FORTHE CORRESPONDING MONOFUNCTIONAL HUMAN PROTEINS. FIFTH, THEEFFECTIVENESS OF EACH COMPOUND WILL BE EXAMINED BY IN VITRO KINETIC ANALYSIS. SUCCESSFUL COMPOUNDS WILL OFFER AN APPROACH TO CONTROLLING MALARIA, A DISEASE WITH AN INCIDENCEOF APPROXIMATELY 200 MILLION CASES WITH GREATER THAN A MILLION FATALITIES PER YEAR. PHASE I WILL BE CONCERNED DOMINANTLY WITH THE OVERPRODUCTION AND PURIFICATION OF THE ENZYME AND SUBSEQUENT CRYSTALLIZATION.