Deciphering key amino acid residues of photosynthetic NADP-malic enzyme

ROSARIO LUNARI; FLORIAN HABERKORN; FERNANDO ZAMARREÑO; MARCELO COSTABEL; MEIKE HÜDIG; VERÓNICA G. MAURINO; CLARISA E. ÁLVAREZ

Córdoba

Congreso; XVII Reunión Anual y X Taller de la Asociación Argentina de Cristalografía; 2022

Asociación Argentina de Cristalografía

Structure-function studies contribute to deciphering how small modifications in the primary structure couldintroduce desirable characteristics into enzymes without affecting its overall function. Malic enzyme (ME) is an enzyme that catalyzes the oxidative decarboxylation of malate to pyruvate, NAD(P)H, and CO2 in the presence of a divalent metal ion. The ME family is made up of evolutionarily related enzymes, with biochemical and structural versatility allows them to participate in a variety of metabolic processes [1]. In this context, we are carrying out structural kinetic characterization of MEs with the potential to be modified for the benefit of generating a higher yield in species with agronomic importance. In relation to this, we recently gained novel information provided by the crystal structural analysis of the photosynthetic NADPME of maize and sorghum which have specific adapted characteristics in comparison to their nophotosynthetic counterpart. We found that the tetrameric photosynthetic have a key E339 residue which iscritical for its malate pH dependent regulation [2]. This residue is located in a pocket that could function as an allosteric site or a site that behaves like a “switch” that triggers this regulation. In order to better understand this mechanism, we performed different synergic structural analysis (molecular dockings, molecular dynamics, protein electrostatics, fluorescence and thermal shift) complemented with kinetics and phylogenetic studies that allow us to found new candidates residues. All these results could provide information to perform direct rational design mutagenesis in order to have optimized proteins to try to improve the photosynthetic efficiency of agronomic crops that has not yet reached their maximum potential.1. C.E. Alvarez, M. Saigo, E. Margarit, C.S. Andreo, M.F. Drincovich, “Kinetics and functional diversityamong the five members of the NADP-malic enzyme family from Zea mays, a C4 species”,Photoshythesis Research 115(1), (2013) 65.2. C.E. Alvarez, A. Bovdilova, A Höppner, C-C Wolff, M. Saigo, F. Trajtenberg, T. Zhang, A. Buschiazzo,L. Nagel-Steger, M.F. Drincovich, M.J. Lercher, V.G. Maurino, “Molecular adaptations of NADP-malicenzyme for its function in C4 photosynthesis in grasses”, Nature Plants. 5 (2019), 755.