CLONING AND CHARACTERIZATION OF A CDNA-ENCODING PHOSPHOFRUCTOKINASE FROM SCHISTOSOMA-MANSONI

Schistosoma mansoni, a human parasitic worm, depends on anaerobic glycolysis as the main source of energy. Phosphofructokinase (ATP: D-fructose-6-phosphase 1-phosphotransferase, EC 2.7.1.11; PFK) limits the rate of glycolysis in these organisms and it has been found to be a target for some antischistosomal agents. A cDNA clone from this parasite has been isolated and characterized. The cDNA is 3046 base pairs long, contains an open reading frame of 2346 bp and codes for a deducted protein of 781 amino acids. The putative protein encoded by the clone has an exact match with the human muscle PFK of 58% and a 73% match when conserved amino acid substitutions are considered. ATP and Fructose-6-P sites have been identified by crystallographic data in the Escherichia coli and Bacillus stearothermophilus PFKs. There is excellent homology between those PFKs and the schistosome PFK at those sites. The PFK-coding cDNA was expressed in insect cells and was shown to ben enzymatically active. Western blot analysis of the recombinant protein in cell extracts gave a positive band with the expected molecular weight of 86 kDa.