Structure determination of the bacteriophage phiX174.

Abstract

The structure of the single-stranded DNA phage phiX174 has been determined to 3.4 A resolution. The crystal space group was P2(1) with one icosahedral particle per asymmetric unit, giving 60-fold noncrystallographic redundancy. Oscillation diffraction photographs were collected using synchrotron radiation at various wavelengths. The particle orientations in the unit cell were determined with a rotation function. Because cowpea mosaic virus has a similar external envelope to phiX174, it was used as a search model to find the approximately positions of the phiX174 particles in the unit cell relative to the crystallographic symmetry axes. An initial phase set to 12 A resolution was then based on the cowpea mosaic virus atomic structure. These phases were improved by 20 cycles of real-space molecular replacement averaging. The phase information was gradually extended to 3.4 A resolution by molecular replacement electron density averaging. One reciprocal lattice point was used for each extension followed by four cycles of averaging. The unusual particle capsid, with its 12 pentameric spikes, required the careful determination of a precise molecular envelope. This was redetermined at regular intervals, as was the particle center. The resultant electron density map was readily interpreted in terms of the F, G and J polypeptides in the capsid. A difference electron density map between full and partially empty particles showed some ordered DNA structure.