title: A novel enzyme with spermine oxidase properties in bovine liver mitochondria: identification and kinetic characterization.
abstract: The uptake of spermine into mammalian mitochondria indicated the need to identify its catabolic pathway in these organelles. Bovine liver mitochondria were therefore purified and their capacity for natural polyamine uptake was verified. A kinetic approach was then used to determine the presence of an MDL 72527-sensitive enzyme with spermine oxidase activity in the matrix of bovine liver mitochondria. Western blot analysis of mitochondrial fractions and immunogold electron microscopy observations of purified mitochondria unequivocally confirmed the presence of a protein recognized by anti-spermine oxidase antibodies in the mitochondrial matrix. Preliminary kinetic characterization showed that spermine is the preferred substrate of this enzyme; lower activity was detected with spermidine and acetylated polyamines. Catalytic efficiency comparable to that of spermine was also found for 1-aminododecane. The considerable effect of ionic strength on the Vmax/KM ratio suggested the presence of more than one negatively charged zone inside the active site cavity of this mitochondrial enzyme, which is probably involved in the docking of positively charged substrates. These findings indicate that the bovine liver mitochondrial matrix contains an enzyme belonging to the spermine oxidase class. Because H2O2 is generated by spermine oxidase activity, the possible involvement of the latter as an important signaling transducer under both physiological and pathological conditions should be considered.
authors.affiliation: Department of Molecular Medicine, University of Padova, Via G. Colombo 3, 35131 Padova, Italy.
authors.forename: Emanuela
authors.initials: E
authors.lastname: Bonaiuto
authors.affiliation: Department of Biomedical Sciences, University of Padova, Via G. Colombo 3, 35131 Padova, Italy.
authors.forename: Silvia
authors.initials: S
authors.lastname: Grancara
authors.affiliation: Department of Biomedical Sciences, University of Padova, Via G. Colombo 3, 35131 Padova, Italy.
authors.forename: Pamela
authors.initials: P
authors.lastname: Martinis
authors.affiliation: Department of Technology and Health, Italian Institute of Health, 00161 Roma, Italy.
authors.forename: Annarita
authors.initials: A
authors.lastname: Stringaro
authors.affiliation: Department of Technology and Health, Italian Institute of Health, 00161 Roma, Italy.
authors.forename: Marisa
authors.initials: M
authors.lastname: Colone
authors.affiliation: Istituto Pasteur-Fondazione Cenci Bolognetti, Department of Biochemical Sciences "A. Rossi Fanelli," Sapienza University of Rome and Institute of Biology and Molecular Pathology, Italian Research Council, P.le Aldo Moro 5, 00185 Roma, Italy.
authors.forename: Enzo
authors.initials: E
authors.lastname: Agostinelli
authors.affiliation: Istituto Pasteur-Fondazione Cenci Bolognetti, Department of Biochemical Sciences "A. Rossi Fanelli," Sapienza University of Rome and Institute of Biology and Molecular Pathology, Italian Research Council, P.le Aldo Moro 5, 00185 Roma, Italy.
authors.forename: Alberto
authors.initials: A
authors.lastname: Macone
authors.affiliation: Department of Molecular Science and Nanosystems, Università Ca' Foscari, Dorsoduro 2137, 30123 Venezia, Italy.
authors.forename: Roberto
authors.initials: R
authors.lastname: Stevanato
authors.affiliation: Department of Comparative Biomedicine and Food Science, Polo Agripolis, Viale dell'Università 16, University of Padova, 35020 Legnaro, Italy; Regional Centre for Advanced Technologies and Materials, Department of Physical Chemistry, Palacky University in Olomouc, 17 Listopadu 1192/12, 771 46 Olomouc, Czech Republic.
authors.forename: Fabio
authors.initials: F
authors.lastname: Vianello
authors.affiliation: Department of Biomedical Sciences, University of Padova, Via G. Colombo 3, 35131 Padova, Italy. Electronic address: antonio.toninello@unipd.it.
authors.forename: Antonio
authors.initials: A
authors.lastname: Toninello
authors.affiliation: Department of Molecular Medicine, University of Padova, Via G. Colombo 3, 35131 Padova, Italy; Consorzio Interuniversitario "Istituto Nazionale Biostrutture e Biosistemi," Viale delle medaglie d'Oro 305, 00136 Roma, Italy. Electronic address: marialuisa.dipaolo@unipd.it.
authors.forename: Maria Luisa
authors.initials: ML
authors.lastname: Di Paolo
mesh_terms: D000818:Animals; D020134:Catalytic Domain; D002417:Cattle; D006861:Hydrogen Peroxide / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008930:Mitochondria, Liver / Q000201:enzymology*; D009994:Osmolar Concentration; D010084:Oxidation-Reduction; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D011700:Putrescine / Q000031:analogs & derivatives / Q000737:chemistry; D013095:Spermidine / Q000378:metabolism; D013096:Spermine / Q000378:metabolism*; D055672:Static Electricity; D013379:Substrate Specificity; D000097667:Polyamine Oxidase
chemical_list: D006861:Hydrogen Peroxide; D000587:Oxidoreductases Acting on CH-NH Group Donors; D011700:Putrescine; D013095:Spermidine; D013096:Spermine; D000097667:Polyamine Oxidase; C044445:MDL 72527
keywords: Immunogold; Mitochondria; Polyamine oxidases; Polyamines; Spermine oxidase