Structural metal dependency of the arginase from the human malaria parasite Plasmodium falciparum.
The human malaria parasite Plasmodium falciparum possesses a single gene with high similarity to the metalloproteins arginase and agmatinase. The recombinant protein reveals strict specificity for arginine, and it has been proposed that its function in ornithine production is as a precursor for polyamine biosynthesis. The specific activity of the plasmodial arginase was found to be 31 micromol min(-1) mg(-1) protein and the k(cat) was calculated as 96 (s-1) . The Km value for arginine and Ki value for ornithine were determined as 13 mM and 19 mM, respectively. The active arginase is a homotrimer of ca. 160 kDa. Dialysis of the arginase against EDTA results in monomers of approximately 48 kDa; however, the quaternary structure can be restored by addition of Mn 2+ . Mutagenic analyses of all the amino acid residues proposed to be involved in metal binding led to complex dissociation, except for the His-193-Ala mutant, which was also inactive but retained the trimeric structure. Substitution of His-233, which has been suggested to be in charge of proton shuttling within the active site, disrupted the trimeric structure and thereby the activity of the Pf arginase. Northern blot analysis identified a stage-specific expression pattern of the plasmodial arginase in the ring/young trophozoite stage, which guarantees the provision of ornithine for essential polyamine biosynthesis.
Müller
Ingrid B
IB
Department of Biochemistry, Bernhard Nocht Institute for Tropical Medicine, Bernhard-Nocht-Strasse 74, D-20359 Hamburg, Germany.
Walter
Rolf D
RD
Wrenger
Carsten
C
D000595:Amino Acid Sequence; D000818:Animals; D001119:Arginase / Q000737:chemistry*; D007700:Kinetics; D008345:Manganese / Q000737:chemistry; D008670:Metals / Q000737:chemistry*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010963:Plasmodium falciparum / Q000201:enzymology* / Q000254:growth & development; D020836:Protein Structure, Quaternary
D008670:Metals; D008345:Manganese; D001119:Arginase