Characterization of novel plasmepsins from the malaria parasite Plasmodium falcipaurm

Though malaria has been largely eradicated in the United States and Western Europe, approximately 40% of the world's population is at risk for infection. Approximately 300 million cases of acute illness occur every year resulting in the death of more than one million people, over 75% of whom are children under the age of five. Four species of Plasmodium are responsible for malaria infection in humans. Of these four, P. falciparum is the most deadly.;Currently, the best methods for keeping the spread of malaria under control are preventative in nature, i.e. spraying with insecticide to eradicate mosquitoes and utilizing insecticide treated bed nets. Though many drugs are available to treat malaria infection, resistance to these continues to grow, even for the artemisinin-based compounds. As growing resistance to current drug therapies surfaces for all four species, a need for novel drug targets to combat infection has arisen.;Aspartic proteases have long been considered an attractive drug target in malaria and various other diseases. This is due to many factors including: (A) proteases often play a crucial role in development of the disease (B) aspartic proteases are the least abundant protease in the human body, which keeps drug interactions within the body to a minimum and (C) structure-based drug design has produced compounds that have been utilized clinically.;Hemoglobin degradation within the digestive vacuole of the P. falciparum parasite was targeted early-on as an essential step in parasite maturation and subsequently enzymes involved in this process were isolated and characterized. The aspartic proteases found within the digestive vacuole are known as plasmepsins 1, 2, 4 and HAP. Studies have indicated that individually these four enzymes are not essential for parasite growth, prompting us to look for other viable drug targets.;Completion of the P. falciparum genome project in 2002 revealed that the parasite encodes for six plasmepsins in addition to the four known to be found within the digestive vacuole. Three of these, plasmepsins 5, 9, and 10, are expressed during the blood stage of the parasite's life cycle and might prove to be novel drug targets. In our laboratory we have successfully expressed and have observed catalytic activity for plasmepsins 9 and 10. We have analyzed these proteases utilizing combinatorial library analysis. We have begun testing plasmepsin 9 with protease inhibitors, including a set of the current clinically used HIV-1 protease inhibitors, pepstatin-based compounds from Sergio Romeo, University of Milan, Milan, Italy, and alpha-substituted norstatins from Kristina Orrling, Uppsala University, Uppsala, Sweden. The data and on-going experiments characterizing these novel targets will provide information that can be used for structure-based drug design studies, leading eventually to a novel drug therapy to combat malaria. (Full text of this dissertation may be available via the University of Florida Libraries web site. Please check http://www.uflib.ufl.edu/etd.html)...