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Plasmid-directed assembly of the lipid-containing membrane of bacteriophage phi 6
1Department of Microbiology, Public Health Research Institute, New York, New York 10016.
Abstract:
The nucleocapsid of bacteriophage phi 6 is enveloped within a lipid-containing membrane. The membrane is composed of proteins P3, P6, P9, P10, and P13 and phospholipids. The relationship between membrane protein P9 and morphogenetic protein P12 was studied in the absence of phage infection. cDNA copies of genes 9 and 12 were expressed on plasmids in Pseudomonas syringae pv. phaseolicola. Immunoblotting demonstrated the presence of protein P9 in strains carrying both gene 9 and gene 12 but not in strains with gene 9 alone. In the absence of P12, P9 was found to be unstable. Simultaneous synthesis of proteins P9 and P12 led to the formation of a low-density P9 particle having a buoyant density similar to that of precursor structures composed of phospholipid and proteins isolated from phi 6-infected cells. These results are consistent with results of previous genetic experiments suggesting that P9 and P12 are necessary and sufficient for the formation of the phi 6 envelope. Extensions of P9 at the C terminus do not impair particle formation; however, N-terminal extensions or C-terminal deletions that extend into the hydrophobic region of P9 do impair particle formation.
Insights
Bacteriophage phi 6 envelope formation requires proteins P9 and P12. Protein P9 is unstable without P12, and together they form precursor particles essential for the viral envelope.
Area of Science:
- Virology
- Molecular Biology
- Structural Biology
Background:
- Bacteriophage phi 6 possesses a lipid-containing membrane envelope.
- This envelope is composed of specific proteins (P3, P6, P9, P10, P13) and phospholipids.
Purpose of the Study:
- To investigate the relationship between membrane protein P9 and morphogenetic protein P12.
- To determine the roles of P9 and P12 in bacteriophage phi 6 envelope formation.
Main Methods:
- Expression of bacteriophage phi 6 genes 9 and 12 using cDNA on plasmids in Pseudomonas syringae pv. phaseolicola.
- Immunoblotting to detect protein P9 presence.
- Analysis of P9 particle formation and buoyant density.
Main Results:
- Protein P9 requires protein P12 for stability; P9 is unstable in the absence of P12.
- Simultaneous synthesis of P9 and P12 resulted in low-density P9 particles.
- The buoyant density of these P9 particles resembled that of precursor structures from infected cells.
Conclusions:
- Proteins P9 and P12 are necessary and sufficient for bacteriophage phi 6 envelope formation.
- Specific structural features of P9, particularly its hydrophobic region, are critical for particle formation.
- N-terminal extensions or C-terminal deletions affecting the hydrophobic region of P9 disrupt envelope assembly.