ATOMIC-STRUCTURE OF THE DEGRADED PROCAPSID PARTICLE OF THE BACTERIOPHAGE-G4 - INDUCED STRUCTURAL-CHANGES IN THE PRESENCE OF CALCIUM-IONS AND FUNCTIONAL IMPLICATIONS

Bacteriophage G4 and phi X174 are members of the Microviridae family T he degree of similarity of the structural proteins ranges from 66% ide ntity of the F protein to 40% identity of the G protein. The atomic st ructure of the phi X174 virion had previously been determined by X-ray crystallography. Bacteriophage G4 procapsids, consisting of the struc tural proteins F,G, D, B, H, and small traces of J but no DNA, were se t up for crystallization. However, the resultant crystals were of degr aded procapsid particles, which had lost the assembly scaffolding prot eins D and B, resulting in particles that resembled empty virions. The structure of the degraded G4 procapsid has been determined to 3.0 Ang strom resolution. The particles crystallized in the hexagonal space gr oup P6(3)22 with unit cell dimensions a = b = 414.2(5) Angstrom and c = 263.0(3) Angstrom. The diffraction data were collected at the Cornel l High Energy Synchrotron Source (CHESS) on film and image plates usin g oscillation photography Packing considerations indicated there were two particles per unit cell. A self-rotation function confirmed that t he particles were positioned on 32 point group special positions in th e unit cell. Initial phases were calculated to 6 Angstrom resolution, based on the known phi X174 virion model. Phase information was then e xtended in steps to 3.0 Angstrom resolution by molecular replacement e lectron density modification and particle envelope generation. The res ulting electron density map was readily interpretable in terms of the F and G polypeptides, as occur in the mature capsid of phi X174. In a few regions of the electron density map there were inconsistencies bet ween the density and the published amino acid sequence. Redetermining the amino acid sequence confirmed that the density was correct. The r. m.s. deviation between the C-alpha backbone of the mature capsid of ph i X174 and the degraded G4 procapsid was 0.36 Angstrom for the F prote in and 1.38 Angstrom for the G protein. This is consistent with the gr eater conservation of the F protein compared to the G protein sequence s among members of the Microviridae family. Functionally important fea tures between phi X174 and G4 had greater conservation. Calcium ions ( Ca2+) were shown to bind to G4 at a general site located near the icos ahedral 3-fold axis on the F protein capsid, equivalent to sites found previously in phi X174. Binding of Ca2+ also caused the ordering of t he conserved region of the DNA binding protein J, which was present in the degraded procapsid particle in the absence of DNA. (C) 1996 Acade mic Press Limited