Characterization of plastidial thioredoxins from Arabidopsis belonging to the new y-type.
The plant plastidial thioredoxins (Trx) are involved in the light-dependent regulation of many enzymatic activities, owing to their thiol-disulfide interchange activity. Three different types of plastidial Trx have been identified and characterized so far: the m-, f-, and x-types. Recently, a new putative plastidial type, the y-type, was found. In this work the two isoforms of Trx y encoded by the nuclear genome of Arabidopsis (Arabidopsis thaliana) were characterized. The plastidial targeting of Trx y has been established by the expression of a TrxGFP fusion protein. Then both isoforms were produced as recombinant proteins in their putative mature forms and purified to characterize them by a biochemical approach. Their ability to activate two plastidial light-regulated enzymes, NADP-malate dehydrogenase (NADP-MDH) and fructose-1,6-bisphosphatase, was tested. Both Trx y were poor activators of fructose-1,6-bisphosphatase and NADP-MDH; however, a detailed study of the activation of NADP-MDH using site-directed mutants of its regulatory cysteines suggested that Trx y was able to reduce the less negative regulatory disulfide but not the more negative regulatory disulfide. This property probably results from the fact that Trx y has a less negative redox midpoint potential (-337 mV at pH 7.9) than thioredoxins f and m. The y-type Trxs were also the best substrate for the plastidial peroxiredoxin Q. Gene expression analysis showed that Trx y2 was mainly expressed in leaves and induced by light, whereas Trx y1 was mainly expressed in nonphotosynthetic organs, especially in seeds at a stage of major accumulation of storage lipids.
Collin
Valérie
V
Institut de Biotechnologie des Plantes, Unité Mixte de Recherche 8618, Centre National de la Recherche Scientifique, Université Paris-Sud, 91405 Orsay cedex, France.
Lamkemeyer
Petra
P
Miginiac-Maslow
Myroslawa
M
Hirasawa
Masakazu
M
Knaff
David B
DB
Dietz
Karl-Josef
KJ
Issakidis-Bourguet
Emmanuelle
E
D017360:Arabidopsis / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000032:analysis / Q000378:metabolism*; D004789:Enzyme Activation; D020869:Gene Expression Profiling; D018506:Gene Expression Regulation, Plant; D008291:Malate Dehydrogenase / Q000378:metabolism; D050538:Malate Dehydrogenase (NADP+); D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000502:physiology; D018087:Plastids / Q000737:chemistry; D020033:Protein Isoforms / Q000378:metabolism; D013879:Thioredoxins / Q000032:analysis / Q000378:metabolism*; D013997:Time Factors
D029681:Arabidopsis Proteins; D020033:Protein Isoforms; D013879:Thioredoxins; D008291:Malate Dehydrogenase; D050538:Malate Dehydrogenase (NADP+)