title: CtpA, a putative Mycobacterium tuberculosis P-type ATPase, is stimulated by copper (I) in the mycobacterial plasma membrane.
abstract: The transport of heavy-metal ions across the plasma membrane is essential for mycobacterial intracellular survival; in this context, P-type ATPases are pivotal for maintenance of ionic gradients and the plasma membrane homeostasis of mycobacteria. To date, the copper ion transport that is mediated by P-type ATPases in mycobacteria is poorly understood. In this work, the ion-specific activation of CtpA, a putative plasma membrane Mycobacterium tuberculosis P-type ATPase, with different heavy-metal cations was assessed. Mycobacterium smegmatis mc(2)155 cells heterologously expressing the M. tuberculosis ctpA gene displayed an increased tolerance to toxic levels of the Cu(2+) ion (4 mM) compared to control cells, suggesting that CtpA is possibly involved in the copper detoxification of mycobacterial cells. In contrast, the tolerance of M. smegmatis recombinant cells against other heavy-metal divalent cations, such as Co(2+), Mn(2+), Ni(2+) and Zn(2+), was not detected. In addition, the ATPase activity of plasma membrane vesicles that were obtained from M. smegmatis cells expressing CtpA was stimulated by Cu(+) (4.9 nmol of Pi released/mg of protein.min) but not by Cu(2+) ions; therefore, Cu(2+) reduction to Cu(+) inside mycobacterial cells is suggested. Finally, the plasma membrane vesicles of M. smegmatis that were enriched with CtpA exhibited an optimal activity at 37 °C and pH 7.9; the apparent kinetic parameters of the enzyme were a K(1/2) of 4.68 × 10(-2) µM for Cu(+), a Vmax of 10.3 U/mg of protein, and an h value of 1.91.
authors.affiliation: Chemistry Department, Faculty of Sciences, Universidad Nacional de Colombia, Carrera 30 N° 45-03, Ciudad Universitaria, 111321, Bogotá, Colombia.
authors.forename: Andrés
authors.initials: A
authors.lastname: León-Torres
authors.forename: Lorena
authors.initials: L
authors.lastname: Novoa-Aponte
authors.forename: Carlos-Y
authors.initials: CY
authors.lastname: Soto
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects* / Q000201:enzymology*; D003300:Copper / Q000378:metabolism / Q000494:pharmacology*; D020102:Mycobacterium smegmatis / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics
chemical_list: D003300:Copper; D000251:Adenosine Triphosphatases