THE INFLUENCE OF A SURFACE-CHARGE ON THE ELECTRONIC AND STERIC STRUCTURE OF A HIGH-POTENTIAL IRON-SULFUR PROTEIN

The recombinant high-potential iron-sulfur protein (HiPIP) iso-I from Ectothiorhodospira halophila has been mutated at position 68. The alph a C of Val 68 is within a 0.6-nm sphere from the closest iron ion of t he cluster, The valine residue has been replaced by a negatively charg ed glutamate residue (V68E) and by a positively charged lysine residue (V68K), With respect to the recombinant wild-type protein the reducti on potentials of the V68E and V68K variants are -21+/-2 and + 29+/-2 m V respectively (200 mM NaCl, pH 7, 25 degrees C). The solution structu re of the V68E mutant was solved up to a pairwise RMSD of 66 pm for ba ckbone atoms and 138 pm for all heavy atoms. The structure of the vari ant is very similar to that of recombinant wild type, indicating that the observed changes in reduction potentials are largely due to the ef fect of the introduced charges. It is proposed that the valence distri bution within the oxidized iron-sulfur cluster is affected only slight ly by the change in charge at position 68, but consistently with a sim ple electrostatic model.