Atypical protein disulfide isomerases (PDI)

Benjamin Selles (1) , Flavien Zannini (1) , Jérémy Couturier (1) , Jean-Pierre Jacquot (1) , Nicolas Rouhier (1)

1 IAM - Interactions Arbres-Microorganismes

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: ENDOPLASMIC-RETICULUM; BOND FORMATION; THIOREDOXIN REDUCTASE; CRYSTAL-STRUCTURE; ESCHERICHIA-COLI; ACTIVE-SITE; in vitro study; disulfide bond; QUIESCIN-SULFHYDRYL OXIDASE; OXIDATION-REDUCTION PROPERTIES; SULFUR CLUSTER BINDING; SEED-STORAGE PROTEINS

Mots clés: populus; isomerase; étude in vitro; disulfure; liaison disulfure

Domaines: Sciences du Vivant [q-bio]

Benjamin Selles, Flavien Zannini, Jérémy Couturier, Jean-Pierre Jacquot, Nicolas Rouhier. Atypical protein disulfide isomerases (PDI): Comparison of the molecular and catalytic properties of poplar PDI-A and PDI-M with PDI-L1A. PLoS ONE, 2017, 12 (3), pp.e0174753. ⟨10.1371/journal.pone.0174753⟩. ⟨hal-01562315⟩