title: Distribution and biochemical properties of an M1-family aminopeptidase in Plasmodium falciparum indicate a role in vacuolar hemoglobin catabolism.
abstract: Aminopeptidases catalyze N-terminal peptide bond hydrolysis and occupy many diverse roles across all domains of life. Here we present evidence that an M1-family aminopeptidase, PfA-M1, has been recruited to specialized roles in the human malaria parasite Plasmodium falciparum. PfA-M1 is abundant in two subcellular compartments in asexual intraerythrocytic parasites; that is, the food vacuole, where the catabolism of host hemoglobin takes place, and the nucleus. A unique N-terminal extension contributes to the observed dual targeting by providing a signal peptide and putative alternate translation initiation sites. PfA-M1 exists as two major isoforms, a nuclear 120-kDa species and a processed species consisting of a complex of 68- and 35-kDa fragments. PfA-M1 is both stable and active at the acidic pH of the food vacuole lumen. Determination of steady-state kinetic parameters for both aminoacyl-β-naphthylamide and unmodified dipeptide substrates over the pH range 5.0-8.5 reveals that k(cat) is relatively insensitive to pH, whereas K(m) increases at pH values below 6.5. At the pH of the food vacuole lumen (5.0-5.5), the catalytic efficiency of PfA-M1 remains high. Consistent with the kinetic data, the affinity of peptidic competitive inhibitors is diminished at acidic pH. Together, these results support a catalytic role for PfA-M1 in the food vacuole and indicate the importance of evaluating the potency of peptidic inhibitors at physiologically relevant pH values. They also suggest a second, distinct function for this enzyme in the parasite nucleus.
authors.affiliation: Department of Biochemistry, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: Ragheb
authors.forename: Seema
authors.initials: S
authors.lastname: Dalal
authors.forename: Kristin M
authors.initials: KM
authors.lastname: Bompiani
authors.forename: W Keith
authors.initials: WK
authors.lastname: Ray
authors.forename: Michael
authors.initials: M
authors.lastname: Klemba
mesh_terms: D000595:Amino Acid Sequence; D000626:Aminopeptidases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D017931:DNA Primers; D004912:Erythrocytes / Q000469:parasitology; D006454:Hemoglobins / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D010963:Plasmodium falciparum / Q000201:enzymology*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014617:Vacuoles / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D006454:Hemoglobins; D000626:Aminopeptidases