REFINED CRYSTAL-STRUCTURE OF SPINACH FERREDOXIN REDUCTASE AT 1.7 ANGSTROM RESOLUTION - OXIDIZED, REDUCED AND 2'-PHOSPHO-5'-AMP BOUND-STATES

The crystal structure of spinach ferredoxin-NADP(+)-oxidoreductase (FN R), determined by multiple isomorphous replacement at 2.6 Angstrom res olution, has been refined at 1.7 Angstrom resolution to an R-factor of 17.9%. The structure of FNR bound to the competitive inhibitor 2'-pho spho-5'-AMP (P-AMP) has also been refined at 1.7 Angstrom to an R-fact or of 17.4% and dithionite-reduced/P-AMP-bound FNR has been refined at 2.0 Angstrom to an R-factor of 14.9%. The P-AMP-bound structure Mras used to construct a model for the binding of NADP(+). Over 200 solvati on sites were included in each structure, and many of the best defined solvation sites stabilize buried turns. A bulk solvent correction obv iated the need for a low-resolution data cutoff. An acidic side-chain likely to be responsible for the low pH requirement for crystallizatio n has been identified. Three large networks of the hydrophobic side-ch ains help define the FNR structure. One of these contains a large cavi ty far from the active site, which coincides with the lone site of seq uence heterogeneity in FNR, and may provide a site for membrane attach ment. The reduced structure shows that Ser96 moves toward atom N-5 of FAD and a water molecule moves toward atom N-1 of FAD, while the flavi n moiety remains planar. Possible sources of a proton that must be pic ked up upon reduction are discussed.