Plant-like phosphofructokinase from Plasmodium falciparum belongs to a novel class of ATP-dependent enzymes.
Malaria parasite-infected erythrocytes exhibit enhanced glucose utilisation and 6-phospho-1-fructokinase (PFK) is a key enzyme in glycolysis. Here we present the characterisation of PFK from the human malaria parasite Plasmodium falciparum. Of the two putative PFK genes on chromosome 9 (PfPFK9) and 11 (PfPFK11), only the PfPFK9 gene appeared to possess all the catalytic features appropriate for PFK activity. The deduced PfPFK proteins contain domains homologous to the plant-like pyrophosphate (PPi)-dependent PFK beta and alpha subunits, which are quite different from the human erythrocyte PFK protein. The PfPFK9 gene beta and alpha regions were cloned and expressed as His(6)- and GST-tagged proteins in Escherichia coli. Complementation of PFK-deficient E. coli and activity analysis of purified recombinant proteins confirmed that PfPFK9beta possessed catalytic activity. Monoclonal antibodies against the recombinant beta protein confirmed that the PfPFK9 protein has beta and alpha domains fused into a 200 kDa protein, as opposed to the independent subunits found in plants. Despite an overall structural similarity to plant PPi-PFKs, the recombinant protein and the parasite extract exhibited only ATP-dependent enzyme activity, and none with PPi. Unlike host PFK, the Plasmodium PFK was insensitive to fructose-2,6-bisphosphate (F-2,6-bP), phosphoenolpyruvate (PEP) and citrate. A comparison of the deduced PFK proteins from several protozoan PFK genome databases implicates a unique class of ATP-dependent PFK present amongst the apicomplexan protozoans.
Mony
Binny M
BM
Department of Biological Sciences, Tata Institute of Fundamental Research, Homi Bhabha Road, Mumbai 400 005, Maharashtra, India. binny@tifr.res.in
Mehta
Monika
M
Jarori
Gotam K
GK
Sharma
Shobhona
S
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D006019:Glycolysis / Q000235:genetics*; D006801:Humans; D010732:Phosphofructokinase-1 / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D010940:Plant Proteins / Q000235:genetics; D010944:Plants / Q000235:genetics / Q000378:metabolism; D010963:Plasmodium falciparum / Q000201:enzymology* / Q000235:genetics; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism
D010940:Plant Proteins; D015800:Protozoan Proteins; D000255:Adenosine Triphosphate; D010732:Phosphofructokinase-1