The structure of the bacteriophage phi X174 was examined in a 2.7 A resolution map and refined, using 6.0 A to 3.0 A resolution data with F > or = 5 sigma (F). The final R-factor was 20.9% and the root-mean-square deviation from idealized bond lengths was 0.021 A. The Hendrickson-Konnert refinement was restrained by the phases derived from the molecular replacement icosahedral averaging procedure. The mature phage capsid consists of 60 copies of the F protein with 426 amino acids, the G protein with 175 amino acids and the J protein with 37 amino acids, as well as 12 copies of the H protein with 328 amino acids. The entire polypeptide chain of the F and G protein, all but the first N-terminal residue of the J protein, and 178 solvent molecules were included in the refinement calculations. The secondary structural features of the F, G and J proteins and their interactions with each other are described. The majority of the protein-protein interactions are between the icosahedral 5-fold related interfaces of the F and of the G proteins. These pentameric units of the F and G proteins form the 9S and 6S assembly intermediates, respectively. The J protein lacks any secondary structure and acts as a linking arm between the icosahedral 5-fold related F proteins. Water molecules were introduced only after phase extension to 2.7 A resolution had been completed. The F protein is associated with lower "thermal" parameters and exhibits greater water order in its environment than the G and J proteins. The largest thermal parameters occur in residues on the viral surface. The solvent contributes to the interactions between the proteins. There is an interface of solvent molecules between the F and the G pentamers which stabilizes the pentameric G protein spikes in a crater centered at each of the icosahedral 5-fold vertices of the F protein capsid. Sequence alignments of the F, G and J amino acid sequences for the homologous bacteriophages G4, alpha 3, phi K and phi X174 with respect to the phi X174 structure demonstrated the conservation of functionally important residues on the viral surface.