Abstract

H2O2 inhibits the [3H]carnitine/carnitine antiport catalysed by the mitochondrial carnitine/acylcarnitinetransporter reconstituted in proteoliposomes. The inhibition was reversed by dithioerythritol, N-acetylcysteineand L-cysteine. Inhibition time-dependence revealed a faster and a slower reaction stages withorders of reaction of 1.0 and 1.9, respectively. Inhibition was tested on mutants in which one or more ofthe six Cys residues had been substituted with Ser or with Val. The four replacement mutant C23S/C58S/C89S/C283S containing C136 and C155 was inhibited as the wild-type. Mutants C23V/C58V/C155V/C89S/C283S and C23V/C58V/C136V/C89S/C283S containing only C136 or C155, respectively, were inhibited ata much lower extent respect to the wild-type, while the mutant C136S/C155S in which the two Cys weresubstituted and the C-less protein were virtually insensitive to inhibition. DTE reversed the inhibition ofthe H2O2 sensitive proteins except that in the case of the mutants containing only C136 or C155 afterlong time of incubation with H2O2. The IC50 values obtained by dose-response curves of H2O2 inhibitionwere 0.17 mM for the wild-type, 0.39 mM for the four replacement mutant containing C136 and C155,2.23 or 1.8 mM in the five replacement mutants containing the single C136 or C155, respectively.Carnitine and acetylcarnitine protected the protein from the inhibition by H2O2. Inhibition kineticsshowed a competitive behaviour of H2O2 respect to carnitine. All the data concur to demonstrate thatH2O2 interacts with C136 and C155 and completely inactivates the transporter by inducing the formationof a disulphide.

Autore Pugliese

• Tonazzi Annamaria

Tutti gli autori

• A. Tonazzi; L. Console; C. Indiveri

Titolo volume/Rivista

Chemico-biological interactions

Anno di pubblicazione

2013

ISSN

0009-2797

ISBN

Non Disponibile

Numero di citazioni Wos

Nessuna citazione

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Numero di citazioni Scopus

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Settori ERC

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Codici ASJC

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