Atypical protein disulfide isomerases (PDI): Comparison of the molecular and catalytic properties of poplar PDI-A and PDI-M with PDI-L1A.
Fiche publication
Date publication
mars 2017
Journal
PloS one
Auteurs
Membres identifiés du Cancéropôle Est :
Dr SELLES Benjamin
Tous les auteurs :
Selles B, Zannini F, Couturier J, Jacquot JP, Rouhier N
Lien Pubmed
Résumé
Protein disulfide isomerases are overwhelmingly multi-modular redox catalysts able to perform the formation, reduction or isomerisation of disulfide bonds. We present here the biochemical characterization of three different poplar PDI isoforms. PDI-A is characterized by a single catalytic Trx module, the so-called a domain, whereas PDI-L1a and PDI-M display an a-b-b'-a' and a°-a-b organisation respectively. Their activities have been tested in vitro using purified recombinant proteins and a series of model substrates as insulin, NADPH thioredoxin reductase, NADP malate dehydrogenase (NADP-MDH), peroxiredoxins or RNase A. We demonstrated that PDI-A exhibited none of the usually reported activities, although the cysteines of the WCKHC active site signature are able to form a disulfide with a redox midpoint potential of -170 mV at pH 7.0. The fact that it is able to bind a [Fe2S2] cluster upon Escherichia coli expression and anaerobic purification might indicate that it does not have a function in dithiol-disulfide exchange reactions. The two other proteins were able to catalyze oxidation or reduction reactions, PDI-L1a being more efficient in most cases, except that it was unable to activate the non-physiological substrate NADP-MDH, in contrast to PDI-M. To further evaluate the contribution of the catalytic domains of PDI-M, the dicysteinic motifs have been independently mutated in each a domain. The results indicated that the two a domains seem interconnected and that the a° module preferentially catalyzed oxidation reactions whereas the a module catalyzed reduction reactions, in line with the respective redox potentials of -170 mV and -190 mV at pH 7.0. Overall, these in vitro results illustrate that the number and position of a and b domains influence the redox properties and substrate recognition (both electron donors and acceptors) of PDI which contributes to understand why this protein family expanded along evolution.
Mots clés
Amino Acid Sequence, Catalysis, Dithionitrobenzoic Acid, metabolism, Humans, Hydrogen-Ion Concentration, Insulin, metabolism, Iron, metabolism, Malate Dehydrogenase (NADP+), chemistry, Mutagenesis, Site-Directed, Oxidoreductases, chemistry, Peroxiredoxins, metabolism, Protein Disulfide-Isomerases, chemistry, Protein Isoforms, chemistry, Sequence Homology, Amino Acid, Sulfides, metabolism, Thioredoxin-Disulfide Reductase, chemistry
Référence
PLoS One. 2017 03 31;12(3):e0174753