Purification, crystallization and preliminary X-ray diffraction studies of the bacteriophage phi29 connector particle.

ABSTRACT

The connector or portal particle from double-stranded DNA bacteriophage phi29 has been crystallized. This structure, which connects the head of the virus with the tail and plays a central role in prohead assembly and DNA packaging and translocation, is formed by 12 subunits of the p10 protein and has a molecular weight of 430 kDa. The connector structure was proteolysed with endoproteinase Glu-C from Staphylococcus aureus V8, which removes 13 and 18 amino acids from the amino- and carboxy-terminal regions of the p10 protein, respectively. Two crystal forms were grown from drops containing an alcohol solution and paraffin oil. Crystals of form I are monoclinic, space group C2 with cell dimensions a = 416.86 A, b = 227.62 A, c = 236.68 A and beta = 96.3 degrees and contain four connector particles per asymmetric unit. Crystals of form II are tetragonal, space group P4(2)2(1)2 with cell dimensions a = b = 170.2 A, c = 156.9 A and contain half a particle per asymmetric unit. X-ray diffraction data from both native crystal forms have been collected to 6.0 and 3.2 A respectively, using synchrotron radiation. Crystals of form II are likely to have the same packing arrangement as the two-dimensional crystals analyzed previously by electron microscopy.