Effect of zinc and calcium ions on the rat kidney membrane-bound form of dipeptidyl peptidase IV |
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Authors: | Hansel Gómez Mae Chappé Pedro A Valiente Tirso Pons María de Los Angeles Chávez Jean-Louis Charli Isel Pascual |
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Institution: | 1. Centro de Estudios de Proteínas (CEP), Facultad de Biología, Universidad de la Habana, Calle 25 No. 455, Vedado, La Habana, 10400, Cuba 2. Departament de Química, Universitat Autònoma de Barcelona, 08193, Bellaterra, Barcelona, Spain 3. Structural Biology and Biocomputing Programme, Spanish National Cancer Research Centre (CNIO), C/Melchor Fernández Almagro 3, Madrid, E-28029, Spain 4. Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México (UNAM), Cuernavaca, Morelos, 62210, México
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Abstract: | Dipeptidyl peptidase IV (DPP-IV) is an ectopeptidase with many roles, and a target of therapies for different pathologies. Zinc and calcium produce mixed inhibition of porcine DPP-IV activity. To investigate whether these results may be generalized to mammalian DPP-IV orthologues, we purified the intact membrane-bound form from rat kidney. Rat DPP-IV hydrolysed Gly-Pro-p-nitroanilide with an average Vmax of 0.86±0.01 μmol min–1mL–1 and KM of 76±6 μM. The enzyme was inhibited by the DPP-IV family inhibitor l-threo-Ile-thiazolidide (Ki=64.0±0.53 nM), competitively inhibited by bacitracin (Ki=0.16±0.01 mM) and bestatin (Ki=0.23±0.02 mM), and irreversibly inhibited by TLCK (IC50 value of 1.20±0.11 mM). The enzyme was also inhibited by divalent ions like Zn2+ and Ca2+, for which a mixed inhibition mechanism was observed (Ki values of the competitive component: 0.15±0.01 mM and 50.0±1.05 mM, respectively). According to bioinformatic tools, Ca2+ ions preferentially bound to the β-propeller domain of the rat and human enzymes, while Zn2+ ions to the α-β hydrolase domain; the binding sites were essentially the same that were previously reported for the porcine DPP-IV. These data suggest that the cationic susceptibility of mammalian DPP-IV orthologues involves conserved mechanisms. |
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