Structure of the azurin mutant Phe114Ala from Pseudomonas aeruginosa at 2.6 Å resolution

The crystal structure of azurin mutant Phe114Ala from Pseudomonas aeruginosa has been solved by molecular replacement. The final crystallographic R value is 0.185 for 9832 reflections to a resolution of 2.6 Å. The root-mean-square deviation for main-chain atom positions is 0.020 Å between the four independent monomers in the asymmetric unit. The mutant Ala114 crystallized from PEG 4000 in a new crystal form and the crystals are monoclinic, P2₁, a= 51.0, b = 83.6, c= 66.4 Å and β = 110.5°. The four molecules in the asymmetric unit are packed as a dimer of dimers and are related by an approximate twofold axis. The dimer packing and the dimer contact region are very similar to that of the Alcaligenes denitrificans azurin dimer. The mutation was performed at residue Phe114, which exhibits a π-electron overlap with the copper ligand His117, to investigate its suggested role in the electron self-exchange reaction. Removal of steric constrains from the phenylalanine side chain created a somewhat different geometry around the copper site with an increased mobility of His117 resulting in an enlarged Cu—N length which may be responsible for the slight differences obtained in the spectral properties of the mutant versus the wild-type protein.