X-RAY STRUCTURE OF THE FERREDOXIN-NADP(-7119 AT 1.8 ANGSTROM RESOLUTION, AND CRYSTALLOGRAPHIC STUDIES OF NADP(+) BINDING AT 2.25 ANGSTROM RESOLUTION() REDUCTASE FROM THE CYANOBACTERIUM ANABAENA PCC)

The crystal structure of the ferredoxin:NADP(+) reductase (FNR) from t he cyanobacterium Anabaena PCC7119 has been determined at 2.6 Angstrom , resolution by multiple isomorphous replacement and refined using 15. 0 Angstrom to 1.8 Angstrom data, collected at 4 degrees C, to an R-fac tor of 0.172. The model includes 303 residues, the flavin adenine dinu cleotide cofactor (FAD), one sulfate ion located at the putative NADP( +) binding site and 328 water molecule sites. The structure of Anabaen a FNR, including FAD, a network of intrinsic water molecules and a lar ge hydrophobic cavity in the C-terminal domain, resembles that of the spinach enzyme. The major differences concern the additional short alp ha-helix (residues 172 to 177 in Anabaena FNR) and residues Arg100 and Arg233 which binds NADP(+) instead of Lys 116 and Lys244 in the spina ch enzyme. Crystals of a complex of Anabaena FNR with NADP(+) were obt ained. The model of the complex has been refined using 15 Angstrom to 2.25 Angstrom X-ray data, collected at -170 degrees C, to an X-factor of 0.186. This model includes 295 residues, FAD, the full NADP(+) (wit h an occupancy of 0.8) and 444 water molecules. The 2'-5' adenine moie ty of NADP(+) binds to the protein as 2'-phospho-5'-AMP to the spinach FNR, The nicotinamide moiety is turned towards the surface of the pro tein instead of stacking onto the FAD isoanoxazine ring as would be re quired for hydride transfer. The model of the complex agrees with prev ious biochemical studies as residues Arg100 and Arg233 are involved in NADP(+) binding and residues Arg77, Lys53 and Lys294, located on the FAD side of the Enzyme, remain free to interact with ferredoxin and fl avodoxin, the physiological partners of ferredoxin:NADP(+) reductase. (C) 1996 Academic Press Limited