Light-activation of NADP-malate dehydrogenase: A highly controlled process for an optimized function

The chloroplastic nicotinamide adenine dinucleotide phosphate-malate dehydrogenase (NADP-MDH) (EC 1.1.1.82), a key enzyme of photosynthetic carbon assimilation of the C4 NADP-malic enzyme type plants, is strictly regulated by light through the ferredoxin-thioredoxin system. It is inactive in the dark, in the oxidized form, and activated in the light by the reduction of specific regulatory disulfides. A site-directed mutagenesis approach allowed localization of the regulatory disulfides in the N- and C-terminal sequence extensions conserved in all the light-regulated chloroplastic malate dehydrogenases. These extensions do not exist in the permanently active NAD-dependent MDHs (EC 1.1.1.37). Biochemical characterization of the mutants and elimination of negative charges at the C-terminus provided evidence for auto-inhibition of the oxidized enzyme by its C-terminal end through interaction with the active site and showed that the more compact structure of the oxidized dimer was linked to the presence of the N-terminal disulfide. The recently published 3-dimensional structures of the oxidized enzyme confirmed the location of the regulatory disulfides and fully support the auto-inhibition hypothesis. Indeed, the C-terminus is trapped inside the active site, interacting with active-site residues, and the N-termini are inserted at the dimer contact area where they are bound by hydrophobic interactions with both subunits. The physiological function of such complex regulation is discussed.