title: Membrane potential regulates mitochondrial ATP-diphosphohydrolase activity but is not involved in progesterone biosynthesis in human syncytiotrophoblast cells.
abstract: ATP-diphosphohydrolase is associated with human syncytiotrophoblast mitochondria. The activity of this enzyme is implicated in the stimulation of oxygen uptake and progesterone synthesis. We reported previously that: (1) the detergent-solubilized ATP-diphosphohydrolase has low substrate specificity, and (2) purine and pyrimidine nucleosides, tri- or diphosphates, are fully dephosphorylated in the presence of calcium or magnesium (Flores-Herrera 1999, 2002). In this study we show that ATP-diphosphohydrolase hydrolyzes first the nucleoside triphosphate to nucleoside diphosphate, and then to nucleotide monophosphate, in the case of all tested nucleotides. The activation energies (Ea) for ATP, GTP, UTP, and CTP were 6.06, 4.10, 6.25, and 5.26 kcal/mol, respectively; for ADP, GDP, UDP, and CDP, they were 4.67, 5.42, 5.43, and 6.22 kcal/mol, respectively. The corresponding Arrhenius plots indicated a single rate-limiting step for each hydrolyzed nucleoside, either tri- or diphosphate. In intact mitochondria, the ADP produced by ATP-diphosphohydrolase activity depolarized the membrane potential (ΔΨm) and stimulated oxygen uptake. Mitochondrial respiration showed the state-3/state-4 transition when ATP was added, suggesting that ATP-diphosphohydrolase and the F1F0-ATP synthase work in conjunction to avoid a futile cycle. Substrate selectivity of the ATP-diphosphohydrolase was modified by ΔΨm (i.e. ATP was preferred over GTP when the inner mitochondrial membrane was energized). In contrast, dissipation of ΔΨm by CCCP produced a loss of substrate specificity and so the ATP-diphosphohydrolase was able to hydrolyze ATP and GTP at the same rate. In intact mitochondria, ATP hydrolysis increased progesterone synthesis as compared with GTP. Although dissipation of ΔΨm by CCCP decreased progesterone synthesis, NADPH production restores steroidogenesis. Overall, our results suggest a novel physiological role for ΔΨm in steroidogenesis.
authors.affiliation: Universidad Nacional Autónoma de México, Facultad de Medicina, Departamento de Bioquímica y Biología Molecular, México City, Mexico. Electronic address: oflores@bq.unam.mx.
authors.forename: Oscar
authors.initials: O
authors.lastname: Flores-Herrera
authors.affiliation: Universidad Nacional Autónoma de México, Facultad de Medicina, Departamento de Bioquímica y Biología Molecular, México City, Mexico.
authors.forename: Sofia
authors.initials: S
authors.lastname: Olvera-Sánchez
authors.affiliation: Universidad Nacional Autónoma de México, Facultad de Medicina, Departamento de Bioquímica y Biología Molecular, México City, Mexico.
authors.forename: Mercedes
authors.initials: M
authors.lastname: Esparza-Perusquía
authors.affiliation: Universidad Nacional Autónoma de México, Facultad de Medicina, Departamento de Bioquímica y Biología Molecular, México City, Mexico.
authors.forename: Juan Pablo
authors.initials: JP
authors.lastname: Pardo
authors.affiliation: Universidad Nacional Autónoma de México, Facultad de Medicina, Departamento de Bioquímica y Biología Molecular, México City, Mexico.
authors.forename: Juan Luis
authors.initials: JL
authors.lastname: Rendón
authors.affiliation: Universidad Nacional Autónoma de México, Facultad de Medicina, Departamento de Bioquímica y Biología Molecular, México City, Mexico.
authors.forename: Guillermo
authors.initials: G
authors.lastname: Mendoza-Hernández
authors.affiliation: Universidad Nacional Autónoma de México, Facultad de Medicina, Departamento de Bioquímica y Biología Molecular, México City, Mexico.
authors.forename: Federico
authors.initials: F
authors.lastname: Martínez
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D015703:Antigens, CD; D001081:Apyrase / Q000378:metabolism*; D002384:Catalysis; D006801:Humans; D053078:Membrane Potential, Mitochondrial; D008928:Mitochondria / Q000201:enzymology*; D011374:Progesterone / Q000096:biosynthesis*; D013379:Substrate Specificity; D014327:Trophoblasts / Q000378:metabolism*
chemical_list: D015703:Antigens, CD; D011374:Progesterone; D000255:Adenosine Triphosphate; D001081:Apyrase; C067869:CD39 antigen
keywords: ATP hydrolysis; ATP-diphosphohydrolase; Mitochondrial bioenergetics; Placental mitochondria; Progesterone synthesis