One of the potential benefits of comparative biochemical studies of parasites is the identification of metabolic deficiencies in the latter and the exploitation of these vulnerabilities for antiparasitic chemotherapy. We have recently verified the absence of de novo synthesis of purine nucleotides in Schistosoma mansoni and proceeded to delineate the purine salvage pathways in this organism. Our results pointed to hypoxanthine-guanine phosphoribosyl transferase (HGPRT) as one of the pivotal enzymes in the purine salvage by S. mansoni; an effective inhibition of this enzyme would lead to death of the parasite. We have since purified this enzyme to apparent homogeneity, partially characterized it and found many of its properties differ from those of the mammalian HGPRT; thus raising the possibility of selective inhibition of the parasite HGPRT. Our future research plan calls for; (1) searching for specific and potent inhibitors of S. mansoni HGPRT and pursuing their possible use as antischistosomal agents; (2) cloning the c-DNA encoding S. mansoni HGPRT and comparing its restriction map and nucleotide sequences with the known amino acid sequence of mammalian HGPRT for possible differences in protein structures between the two enzymes; (3) cloning the full-length c-DNA of S. mansoni HGPRT. This will be done either by introducing a specially constructed expression vector plasmid into an Escherichia coli #165 strain deleted in xanthine-guanine phosphoribosyl transferase (XGPRT) for expression; or by introducing a c-DNA library of S. mansoni constructed in retroviral LTR chimeric plasmids into a mammalian HGPRT cell line and selecting for the expression of S. mansoni HGPRT in HAT medium. The native S. mansoni HGPRT thus produced in larger quantities by these methods will facilitate more thorough investigations on the kinetic and structural of the enzyme; (4) cloning the genomic DNA encoding S. mansoni HGPRT and comparing its full size, restriction pattern and nucleotide sequences with the full-length S. mansoni HGPRT c-DNA to search for possible introns in the gene structure and any regulatory segments flanking the gene in order to better understand the genetic regulation of S. mansoni HGPRT. We have also identified a tubercidin kinase in S. mansoni which has a unique substrate specificity and is the target of the potent antischistosomal activity of tubercidin. We plan to further characterize this enzyme as a potential target for antischistosomal chemotherapy.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Unknown (R22)
Project #
5R22AI024011-03
Application #
3444941
Study Section
Bacteriology and Mycology Subcommittee 1 (BM)
Project Start
1986-08-01
Project End
1989-07-31
Budget Start
1988-08-01
Budget End
1989-07-31
Support Year
3
Fiscal Year
1988
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Type
Schools of Pharmacy
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
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Eakin, A E; Nieves-Alicea, R; Tosado-Acevedo, R et al. (1995) Comparative complement selection in bacteria enables screening for lead compounds targeted to a purine salvage enzyme of parasites. Antimicrob Agents Chemother 39:620-5
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Craig 3rd, S P; McKerrow, J H; Newport, G R et al. (1988) Analysis of cDNA encoding the hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) of Schistosoma mansoni;a putative target for chemotherapy. Nucleic Acids Res 16:7087-101