tailieunhanh - Báo cáo khoa học: Thermodynamic characterization of substrate and inhibitor binding to Trypanosoma brucei 6-phosphogluconate dehydrogenase

6-Phosphogluconate dehydrogenase is a potential target for new drugs against African trypanosomiasis. Phosphorylated aldonic acids are strong inhibitors of 6-phosphogluconate dehydrogenase, and 4-phospho-d-erythro-nate (4PE) and 4-phospho-d-erythronohydroxamate are two of the stron-gest inhibitors of theTrypanosoma bruceienzyme. | ễFEBS Journal Thermodynamic characterization of substrate and inhibitor binding to Trypanosoma brucei 6-phosphogluconate dehydrogenase Katy Montin Carlo Cervellati Franco Dallocchio and Stefania Hanau Dipartimento di Biochimica e Biologia Molecolare Universita di Ferrara Italy Keywords enzyme inhibitors isothermal titration calorimetry 6-phosphogluconate dehydrogenase transition state analogues Trypanosoma brucei Correspondence S. Hanau Dipartimento di Biochimica e Biologia Molecolare Universita di Ferrara Via L. Borsari 46 44100 Ferrara Italy Fax 39 0532202723 Tel 39 0532455443 E-mail hns@ Received 20 July 2007 revised 19 October 2007 accepted 23 October 2007 doi 6-Phosphogluconate dehydrogenase is a potential target for new drugs against African trypanosomiasis. Phosphorylated aldonic acids are strong inhibitors of 6-phosphogluconate dehydrogenase and 4-phospho-D-erythro-nate 4PE and 4-phospho-D-erythronohydroxamate are two of the strongest inhibitors of the Trypanosoma brucei enzyme. Binding of the substrate 6-phospho-D-gluconate 6PG the inhibitors 5-phospho-D-ribonate 5PR and 4PE and the coenzymes NADP NADPH and NADP analogue 3-amino-pyridine adenine dinucleotide phosphate to 6-phospho-D-gluconate dehydrogenase from T. brucei was studied using isothermal titration calorimetry. Binding of the substrate Kd 5 pM and its analogues Kd pM and Kd pM for 5PR and 4PE respectively is entropy driven whereas binding of the coenzymes is enthalpy driven. Oxidized coenzyme and its analogue but not reduced coenzyme display a half-site reactivity in the ternary complex with the substrate or inhibitors. Binding of 6PG and 5PR poorly affects the dissociation constant of the coenzymes whereas binding of 4PE decreases the dissociation constant of the coenzymes by two orders of magnitude. In a similar manner the Kd value of 4PE decreases by two orders of magnitude in the presence of the coenzymes. The results suggest that 5PR acts as