Modulation, via protein-protein interactions, of glyceraldehyde-3-phosphate dehydrogenase activity through redox phosphoribulokinase regulation.
The activity of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) embedded in the phosphoribulokinase (PRK).GAPDH.CP12 complex was increased 2-3-fold by reducing agents. This occurred by interaction with PRK as the cysteinyl sulfhydryls (4 SH/subunit) of GAPDH within the complex were unchanged whatever the redox state of the complex. But isolated GAPDH was not activated. Alkylation plus mass spectrometry also showed that PRK had one disulfide bridge and three SH groups per monomer in the active oxidized complex. Reduction disrupted this disulfide bridge to give 2 more SH groups and a much more active enzyme. We assessed the kinetics and dynamics of the interactions between PRK and GAPDH/CP12 using biosensors to measure complex formation in real time. The apparent equilibrium binding constant for GAPDH/CP12 and PRK was 14 +/- 1.6 nm for oxidized PRK and 62 +/- 10 nm for reduced PRK. These interactions were neither pH- nor temperature-dependent. Thus, the dynamics of PRK.GAPDH.CP12 complex formation and GAPDH activity are modulated by the redox state of PRK.
Lebreton
Sandrine
S
Institut Jacques Monod, UMR 7592 CNRS, Universités Paris VI-VII, 2 place Jussieu, 75251 Paris cedex 05, France.
Graciet
Emmanuelle
E
Gontero
Brigitte
B
D000818:Animals; D015374:Biosensing Techniques; D016825:Chlamydomonas reinhardtii; D002736:Chloroplasts / Q000201:enzymology*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism*; D016877:Oxidants / Q000494:pharmacology; D010084:Oxidation-Reduction; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000378:metabolism; D020349:Surface Plasmon Resonance; D017186:Titrimetry
D016877:Oxidants; D010940:Plant Proteins; D013438:Sulfhydryl Compounds; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D017853:Phosphotransferases (Alcohol Group Acceptor); C022390:phosphoribulokinase