Crystallization and preliminary X-ray diffraction analysis of bacteriophage ϕ12 packaging factor P7

Abstract

Bacteriophage varphi12 protein P7 is a structural component of the polymerase complex and ensures stable packaging of the genomic RNA. varphi12 P7 has been cloned, purified and crystallized. Crystals belong to space group P3(2)21, with unit-cell parameters a = 75.7, b = 75.7, c = 45.2 A, alpha = 90, beta = 90, gamma = 120 degrees , and diffract beyond 2.0 A. Multiple anomalous dispersion data have been collected from crystals of selenomethionylated P7. Mass spectroscopy showed proteolysis of the crystallized protein and a truncated form, P7DeltaC, gave crystals of similar morphology. Cross-linking experiments implicated the N-terminal domain of P7 as being essential for dimerization.

Structure and Dynamics of the P7 Protein from the Bacteriophage ϕ12

“…These intense resonances were not seen in a construct of P7 (P7ΔC, 1–129) lacking the last 40 residues from the C-terminal tail (Figure 1(a)). This is consistent with previous suggestions that the P7 protein in the ϕ6 20 and ϕ12 21 cystoviruses possess flexible C-terminal tails.…”

“…The P7 core (P7ΔC) crystallized in a P3 2 21 space-group, the same space group and similar unit cell parameters as reported in previous preliminary crystallization studies 21, and we solved its structure to a resolution of 1.8 Å using single wavelength anomalous diffraction (SAD). Details of the data collection, structure refinement and structural statistics are provided in Table 1.…”

Roles of the Minor Capsid Protein P7 in the Assembly and Replication of Double-Stranded RNA Bacteriophage ϕ6

“…Purified P7 from a related bacteriophage ϕ12 also easily undergoes proteolytic degradation to produce a C-terminally truncated molecule with 40 C-terminal residues removed, and the truncated polypeptide forms stable dimers. 32 Interestingly, the protease-sensitive loop region, located approximately 40 residues from the C-terminus, and the N-terminal dimerization domain are conserved between the ϕ6 and ϕ12 P7 proteins (Fig. 5a) although the sequence similarity is low (see reference Fig.…”

Hexameric molecular motors: P4 packaging ATPase unravels the mechanism

“…F4 helicases generally form hexameric ring structures, unwind DNA in the 5¢ to 3¢ direction, and often associate physically with DNA primases and function in DNA replication of bacteria and bacteriophages [62]. P4 proteins are hexameric, ssRNA-stimulated NTPases [34,55,56]. f12 P4 exhibits strict specificity for purine nucleotide triphosphates, whereas P4 proteins from the other three bacteriophages hydrolyse both purines and Cell.…”

“…P4 is a hexameric packaging ATPase, structurally related to RecA/F1-ATPase-like motors P4 enzymes from four cystoviruses (f6, f8, f12 and f13) were cloned, expressed and characterized (Table 1) [34,51,[55][56][57][58][59]. P4 proteins belong to the F4 hexameric helicase family, also known as the DnaB-like family [60].…”