title: Kinetic properties and inhibition of the dimeric dUTPase-dUDPase from Campylobacter jejuni.
abstract: The enzyme deoxyuridine 5'-triphosphate nucleotidohydrolase (dUTPase) catalyses the hydrolysis of dUTP to dUMP and PPi thus controlling the incorporation of uracil into DNA genomes. In Campylobacter jejuni dUTPase exhibits structural properties of dimeric proteins characteristic of protozoa of the Kinetoplastidae family. In the present study we perform a kinetic analysis of Campylobacter dUTPase using the continuous spectrophotometric method and show that the enzyme is highly specific for deoxyuridine nucleotides. The Michaelis-Menten constant for dUTP was 0.66 microM while the k(cat) was 12.3 s(- 1). dUDP was also efficiently hydrolysed although the specificity constant, k(cat)/K(m), was five fold lower than for dUTP. The reaction product and the non hydrolysable analogue alpha,beta imido dUDP are potent inhibitors of the enzyme while several analogues of dUMP with substituents at the 3'- and 5'-positions active against trimeric dUTPases, show poor inhibitory activity. Apparent structural and kinetic differences with other eukaryotic dUTPases suggest that the present enzyme might be exploited as a target for new drugs against campylobacteriosis.
authors.affiliation: Instituto de Parastiologia y Biomedicina "Lopez-Neyra". Consejo Superior de Investigaciones Cientificas, Armilla, Granada, Spain.
authors.forename: Juan A
authors.initials: JA
authors.lastname: Musso-Buendía
authors.forename: Antonio E
authors.initials: AE
authors.lastname: Vidal
authors.forename: Ganasan
authors.initials: G
authors.lastname: Kasinthan
authors.forename: Corinne
authors.initials: C
authors.lastname: Nguyen
authors.forename: Juana
authors.initials: J
authors.lastname: Carrero-Lérida
authors.forename: Luis M
authors.initials: LM
authors.lastname: Ruiz-Pérez
authors.forename: Keith
authors.initials: K
authors.lastname: Wilson
authors.forename: Nils Gunnar
authors.initials: NG
authors.lastname: Johansson
authors.forename: Ian H
authors.initials: IH
authors.lastname: Gilbert
authors.forename: Dolores
authors.initials: D
authors.lastname: González-Pacanowska
mesh_terms: D001426:Bacterial Proteins; D016123:Campylobacter jejuni / Q000201:enzymology*; D019281:Dimerization; D006868:Hydrolysis; D007700:Kinetics; D011755:Pyrophosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013379:Substrate Specificity; D014544:Uridine Triphosphate / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D011755:Pyrophosphatases; C023273:uridine diphosphatase; C020020:dUTP pyrophosphatase; D014544:Uridine Triphosphate