Mevinolin is a specific and potent inhibitor of the rate-limiting enzyme of the mevalonate pathway, HMG-CoA reductase. Mevinolin stops cell growth and can block the action of oncogenes, a group of proteins involved in cell proliferation. Mevinolin stops egg production by Schistosoma mansoni and if the drug is administered chronically into infected mice it will eliminate the infection. The drug's action on schistosomes can be abrogated by mevalonate and farnesol a 15 carbon isoprenoid. The major endproducts of the mevalonate pathway, which consist of cholesterol, dolichols, ubiquinones, hema-a juvenile hormones (in insects) and isopentenyl-tRNA cannot abrogate the effects of mevinolin. The recent discovery that certain oncogenes and other G-binding proteins can be posttranslationally modified on their carboxy terminus with farnesyl (C-15) or geranylgeranyl (C-20) and that the biological activity of these modified proteins depends upon their prenylation has raised the question of their role in schistosomes. Schistosome proteins can be labeled with mevalonate and in particular those having a molecular weight of 25 kDa. These 25 kDa mevalonate labeled proteins bind GTP and the lipid attached to them appears to be geranylgeranyl. At least two prenoid:protein transferase enzymes have been characterized and there are endogenous schistosome proteins which these enzymes can prenylate. Farnesyl protein transferase (FPT) selectively labels 46 kDa schistosome proteins while geranylgeranyl protein transferase (GGPT) labels 25 kDa proteins . Schistosome FPT will farnesylate p2l-Hras while the parasite's GGPT will geranylgeranylate a mammalian protein (rab1b) involved in cell trafficking. From this data we have two major objectives, based upon the working hypothesis that prenylated G-binding proteins are vital for parasite function. Our first objective is to purify one of the 25 kDa prenylated proteins and determine how it functions within the schistosome. Our second objective is to characterize and purify schistosome FPT. Classical protein purification methods will be employed to purify and obtain amino acid sequence data on the 25 kDa prenylated protein. Sequence information from this schistosome protein and from highly conserved regions of sequenced eukaryotic G-bind proteins will be used for the construction of probes for probing cDNA libraries or parasite RNA or genomic DNA using PCR. Cloned genes will be sequenced, expressed and purified. Numerous studies on the recombinant 25 kDa protein will attempt to determine its function. Our second objective will focus on the biochemical characteristics of the schistosome's FPT. The proposed research offers us the opportunity to study the function of a novel group of lipid modified schistosome proteins that probably play a vital role in regulating schistosome reproductive processes as well as other vital functions and serve as potential targets for the development of an antischistosomal drug and/or vaccine.
