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UMP kinase from the Gram-positive bacterium Bacillus subtilis is strongly dependent on GTP for optimal activity

Domenii publicaţii > Biologie + Tipuri publicaţii > Articol în revistã ştiinţificã

Autori: Cristina Gagyi, Nadia Bucurenci, Ovidiu Sirbu, Gilles Labesse, Mihaela Ionescu, Augustin Ofiteru, Liliane Assairi, Stéphanie Landais, Antoine Danchin, Octavian Barzu and Anne-Marie Gilles

Editorial: European Journal of Biochemistry, 270, p.3196-3204, 2003.

Rezumat:

The gene encoding Bacillus subtilis UMP kinase (pyrH/smbA) is transcribed in vivo into a functional enzyme, which represents approximately 0.1% of total soluble proteins. The specific activity of the purified enzyme under optimal conditions is 25 units©mg-1 of protein. In the absence of GTP, the activity of B. subtilis enzyme is less than 10% of its maximum activity. Only dGTP and 3′-anthraniloyl-2′-deoxyguanosine-5′-triphosphate (Ant-dGTP) can increase catalysis significantly. Binding of Ant-dGTP to B. subtilis UMP kinase increased the quantum yield of the fluorescent analogue by a factor of more than three. UTP and GTP completely displaced Ant-dGTP, whereas GMP and UMP were ineffective. UTP inhibits UMP kinase of B. subtilis with a lower affinity than that shown towards the Escherichia coli enzyme. Among nucleoside monophosphates, 5-fluoro-UMP (5F-UMP) and 6-aza-UMP were actively phosphorylated by B. subtilis UMP kinase, explaining the cytotoxicity of the corresponding nucleosides towards this bacterium. A structural model of UMP kinase, based on the conservation of the fold of carbamate kinase and N-acetylglutamate kinase (whose crystals were recently resolved), was analysed in the light of physicochemical and kinetic differences between B. subtilis and E. coli enzymes.

Cuvinte cheie: UMP kinaza, B. subtilis, modelare moleculara, activare GTP-dependenta, markeri fluorescenti // UMP kinase; B. subtilis; molecular modelling; GTP activation; fluorescent markers.