Effect of zinc and calcium ions on the rat kidney membrane-bound form of dipeptidyl peptidase IV.
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 meu mol min-1mL-1 and KM of 76 +/- 6 meu M. 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 beta-propeller domain of the rat and human enzymes, while Zn2+ ions to the alpha-beta 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.
Centro de Estudios de Proteinas (CEP), Facultad de Biologia, Universidad de la Habana, Calle 25 No. 455, Vedado, La Habana 10400, Cuba.
Hansel
H
Gómez
Mae
M
Chappé
Pedro A
PA
Valiente
Tirso
T
Pons
María de Los Angeles
Mde L
Chávez
Jean-Louis
JL
Charli
Isel
I
Pascual
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry*; D018819:Dipeptidyl Peptidase 4 / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D007668:Kidney / Q000201:enzymology*; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification; D051381:Rats; D015032:Zinc / Q000737:chemistry*
D008565:Membrane Proteins; D018819:Dipeptidyl Peptidase 4; D015032:Zinc; D002118:Calcium