Modulation Of The Enzyme Catalytic Efficiency By Amino Acid Volume

MUSUMECI, MATIAS M.; RIAL, D; CATALANO DUPUY, D; ARAKAKI, A; CECCARELLI, EDUARDO AUGUSTO

Pinamar, Argentina

Congreso; 10th Congress Panamerican Association for Biochemistry and Molecular Biology; 2005

SAIB - PABMB

Ferredoxin-NADP(H) reductases (FNR) are ubiquitous flavoenzymes that participate in a broad range of redox metabolic pathways. Several structural features of these enzymes remain yet to be explained. FNRs consist of two domains; one involved in the binding of the prosthetic group FAD and the other in the binding of NADP. The residue Y308 in pea FNR is stacked near parallel to the re-face of the flavin and is highly conserved among members of the family. Computing the relative free energy for the lumiflavin-phenol pair with the relative positions found for Y308 in pea FNR we have concluded that this amino acid is constrained against the isoalloxazine. This effect is probably performed by amino acids C266 and L268, which are facing the other side of this tyrosine, forcing it to adopt a more planar orientation with respect to the flavin. Simple and double FNR mutants of amino acids C266 and L268 were obtained and characterized. We observed that reducing amino acid volume decreases the catalytic efficiency without altering the protein structure, probably due to an increase of the Y308 - isoalloxazine interaction. Our results allows to suggest that these amino acids have been evolutively selected by volume and that they participate in the fine tuning of the enzyme efficiency, modulating the interaction of the Y308 with the isoalloxazine.