Ligand-induced heme ruffling and bent NO geometry in ultra-high-resolutionstructures of nitrophorin 4

The nitrophorins are a family of proteins that use ferric heme to transport nitric oxide (NO) from the salivary glands of blood-sucking insects to the ir victims, resulting in vasodilation and reduced blood coagulation. We hav e refined atomic resolution structures of nitrophorin 4 (NP4) from Rhodnius prolixus complexed with NO (1.08 Angstrom) and NH3 (1.15 Angstrom), yieldi ng a highly detailed picture of the iron coordination sphere. In NP4-NO, th e NO nitrogen is coordinated to iron (Fe-N distance = 1.66 Angstrom) and is somewhat bent (Fe-N-O angle = 156 degrees), with bending occurring in the same plane as the proximal histidine ring, The Fe(NO)(heme)(His) coordinati on geometry is unusual but consistent with an Fe(III) oxidation state that is stabilized by a highly ruffled heme. Heme ruffling occurs in both struct ures, apparently due to close contacts between the heme and leucines 123 an d 133, but increases on binding NO even though the steric contacts have not changed. We also report the structure of NP4 in complexes with histamine ( 1.50 Angstrom) and imidazole (1.27 Angstrom). Unexpectedly, two mobile loop s that rearrange to pack against the bound NO in NP4-NO, also rearrange in the NP4-imidazole complex. This conformational change is apparently driven by the nonpolar nature of the NO and imidazole (as bound) ligands. Taken to gether, the desolvation of the NO binding pocket through a change in protei n conformation, and the bending of the NO moiety, possibly through protein- assisted heme ruffling, may lead to a nitrosyl-heme complex that is unusual ly resistant to autoreduction.