Structure of C73G putidaredoxin from Pseudomonas putida

The structure of the C73G mutant of putidaredoxin (Pdx), the Fe 2 S 2 ferredoxin that supplies electrons to cytochrome CYP101 (P450cam) for camphor oxidation, is reported at 1.9 Å resolution in a C 2 crystal form. The structure was solved by single‐wavelength iron anomalous diffraction, which yielded electron density above the 2σ level for over 97% of the non‐H atoms in the protein. The final structure with R  = 0.19 and R free = 0.21 has been deposited in the Protein Data Bank with accession code 1r7s . The C 2 crystal contains three Pdx molecules in the asymmetric unit, giving three independent models of the protein that are very similar (r.m.s.d. < 0.3 Å for the 106 C α atoms). The unusually high solvent fraction of 80% results in comparatively few crystal‐packing artifacts. The structure is briefly compared with the recently reported crystal structures of the C73S and C73S/C85S mutants. In general, the eight independent molecules in the three crystal structures (three in C73G, three in C73S and two in C73S/C85S) are much more similar to each other than to the previously reported NMR structure of wild‐type Pdx in solution. The present findings show a unanimous structure in some regions crucial for electron‐transfer interactions, including the cluster‐binding loop 39–48 and the cytochrome‐interaction region of Asp38 and Trp106. In addition, the Cys45 amide group donates a hydrogen bond to cluster sulfur S1, with Ala46 adopting an Lα conformation, in all three molecules in the crystal.