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Effect of zinc and calcium ions on the rat kidney membrane-bound form of dipeptidyl peptidase IV

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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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Abbreviations

aas:

amino acids

APA:

glutamyl aminopeptidase

APB:

arginyl aminopeptidase

APN:

alanyl aminopeptidase

BSA:

bovine serum albumin

DPP-IV:

dipeptidyl peptidase IV

GIP:

gastric inhibitory peptide

GLP-1:

glucagon-like peptide 1

NEM:

N-ethyl maleimide

P32/98:

l-threo-Ile-thiazolidide

pNA:

p-nitroanilide

TLCK:

tosyl-l-lysine chloromethyl ketone

Xaa:

any common amino acids

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Acknowledgements

This work was funded by grants from the International Foundation for Science (IFS) and the Organization for the Prohibition of Chemical Weapons (OPCW) (F/3276-3), CONACYT-Mexico (61804), CONACYT-Mexico-Cuba-CITMA (2009-2011), and OPCW Internship 2010 to IP.

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Authors and Affiliations

  1. Centro de Estudios de Proteínas (CEP), Facultad de Biología, Universidad de la Habana, Calle 25 No. 455, Vedado, La Habana, 10400, Cuba

    Hansel Gómez, Mae Chappé, Pedro A Valiente, María de Los Angeles Chávez & Isel Pascual

  2. Departament de Química, Universitat Autònoma de Barcelona, 08193, Bellaterra, Barcelona, Spain

    Hansel Gómez

  3. Structural Biology and Biocomputing Programme, Spanish National Cancer Research Centre (CNIO), C/Melchor Fernández Almagro 3, Madrid, E-28029, Spain

    Tirso Pons

  4. Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México (UNAM), Cuernavaca, Morelos, 62210, México

    Jean-Louis Charli

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  1. Hansel Gómez
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  2. Mae Chappé
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  3. Pedro A Valiente
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  5. María de Los Angeles Chávez
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  6. Jean-Louis Charli
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  7. Isel Pascual
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Correspondence to Isel Pascual.

Additional information

Corresponding editor: MARÍA LUZ CÁRDENAS

MS received 16 January 2013; accepted 08 April 2013

Corresponding editor: María Luz Cárdenas

[Gómez H, Chappé M, Valiente P, Pons T, Chávez MLA, Charli J-L and Pascual I 2013 Effect of zinc and calcium ions on the rat kidney membrane-bound form of dipeptidyl peptidase IV. J. Biosci. 38 1–9] DOI 10.1007/s12038-013-9333-8

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Gómez, H., Chappé, M., Valiente, P.A. et al. Effect of zinc and calcium ions on the rat kidney membrane-bound form of dipeptidyl peptidase IV. J Biosci 38, 461–469 (2013). https://doi.org/10.1007/s12038-013-9333-8

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