title: The poplar Phi class glutathione transferase: expression, activity and structure of GSTF1.
abstract: Glutathione transferases (GSTs) constitute a superfamily of enzymes with essential roles in cellular detoxification and secondary metabolism in plants as in other organisms. Several plant GSTs, including those of the Phi class (GSTFs), require a conserved catalytic serine residue to perform glutathione (GSH)-conjugation reactions. Genomic analyses revealed that terrestrial plants have around ten GSTFs, eight in the Populus trichocarpa genome, but their physiological functions and substrates are mostly unknown. Transcript expression analyses showed a predominant expression of all genes both in reproductive (female flowers, fruits, floral buds) and vegetative organs (leaves, petioles). Here, we show that the recombinant poplar GSTF1 (PttGSTF1) possesses peroxidase activity toward cumene hydroperoxide and GSH-conjugation activity toward model substrates such as 2,4-dinitrochlorobenzene, benzyl and phenetyl isothiocyanate, 4-nitrophenyl butyrate and 4-hydroxy-2-nonenal but interestingly not on previously identified GSTF-class substrates. In accordance with analytical gel filtration data, crystal structure of PttGSTF1 showed a canonical dimeric organization with bound GSH or 2-(N-morpholino)ethanesulfonic acid molecules. The structure of these protein-substrate complexes allowed delineating the residues contributing to both the G and H sites that form the active site cavity. In sum, the presence of GSTF1 transcripts and proteins in most poplar organs especially those rich in secondary metabolites such as flowers and fruits, together with its GSH-conjugation activity and its documented stress-responsive expression suggest that its function is associated with the catalytic transformation of metabolites and/or peroxide removal rather than with ligandin properties as previously reported for other GSTFs.
authors.affiliation: Interactions Arbres - Microorganismes, Université de Lorraine, UMR1136 Vandoeuvre-lès-Nancy, France ; INRA, Interactions Arbres - Microorganismes, UMR1136 Champenoux, France.
authors.forename: Henri
authors.initials: H
authors.lastname: Pégeot
authors.affiliation: Faculté des Sciences et Technologies, Université de Lorraine, CRM2, Equipe BioMod, UMR 7036 Vandoeuvre-lès-Nancy, France ; Faculté des Sciences et Technologies, CNRS, CRM2, Equipe BioMod, UMR 7036 Vandoeuvre-lès-Nancy, France.
authors.forename: Cha San
authors.initials: CS
authors.lastname: Koh
authors.affiliation: Interactions Arbres - Microorganismes, Université de Lorraine, UMR1136 Vandoeuvre-lès-Nancy, France ; INRA, Interactions Arbres - Microorganismes, UMR1136 Champenoux, France.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Petre
authors.affiliation: Faculté des Sciences et Technologies, Université de Lorraine, CRM2, Equipe BioMod, UMR 7036 Vandoeuvre-lès-Nancy, France ; Faculté des Sciences et Technologies, CNRS, CRM2, Equipe BioMod, UMR 7036 Vandoeuvre-lès-Nancy, France.
authors.forename: Sandrine
authors.initials: S
authors.lastname: Mathiot
authors.affiliation: Interactions Arbres - Microorganismes, Université de Lorraine, UMR1136 Vandoeuvre-lès-Nancy, France ; INRA, Interactions Arbres - Microorganismes, UMR1136 Champenoux, France.
authors.forename: Sébastien
authors.initials: S
authors.lastname: Duplessis
authors.affiliation: Interactions Arbres - Microorganismes, Université de Lorraine, UMR1136 Vandoeuvre-lès-Nancy, France ; INRA, Interactions Arbres - Microorganismes, UMR1136 Champenoux, France.
authors.forename: Arnaud
authors.initials: A
authors.lastname: Hecker
authors.affiliation: Faculté des Sciences et Technologies, Université de Lorraine, CRM2, Equipe BioMod, UMR 7036 Vandoeuvre-lès-Nancy, France ; Faculté des Sciences et Technologies, CNRS, CRM2, Equipe BioMod, UMR 7036 Vandoeuvre-lès-Nancy, France.
authors.forename: Claude
authors.initials: C
authors.lastname: Didierjean
authors.affiliation: Interactions Arbres - Microorganismes, Université de Lorraine, UMR1136 Vandoeuvre-lès-Nancy, France ; INRA, Interactions Arbres - Microorganismes, UMR1136 Champenoux, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Rouhier
keywords: Populus; crystallography; enzyme characterization; glutathione transferase; protein structure; transcript profiling