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DNA replication and head assembly in bacteriophage T4
Abstract:
Phage DNA was accumulated in cells of E. coli B, infected with the phage T4DtsLB3 (gene 42), without the synthesis of late proteins (in the presence of chloramphenicol). Then (stage II), chloramphenicol was removed and further replication of the phage DNA suppressed with hydroxyurea and by simultaneously raising the temperature to 40 degrees. The media M9 or M9 with 1% amino acid were used; the times of addition of chloramphenicol and the hydroxyurea concentration were also varied. It was also shown that in medium M9, at stage II, chiefly early proteins were synthesized. In the medium containing amino acids, at stage II the following was observed: 1) DNA synthesis was entirely suppressed and a degradation of DNA occurred; 2) both early and late proteins were synthesized, with a predominance of the latter; 3) an assembly of the elements of the phage tails and capsids occurred without the neck and flagellum, and a small number of phage particles were also found; 4) the capsids, isolated in a sucrose density gradient after lysis with chloroform, contained the proteins Palt, P20, P23, P24, several unidentified proteins, and did not contain Pwac, P23, and P22, 5) the yield of viable phage varied from 0.05 to 15% per cell. Thus, the entire morphogenesis of T4 phage can occur without accompanying replication of phage DNA.
Insights
Bacteriophage T4 morphogenesis can occur independently of phage DNA replication. This study demonstrates that essential phage components assemble even when DNA synthesis is inhibited, revealing a novel aspect of viral assembly.
Area of Science:
- Molecular Biology
- Virology
- Microbiology
Background:
- Bacteriophage T4 infection involves DNA replication and synthesis of early and late proteins.
- Viral morphogenesis typically requires coordinated DNA replication and protein synthesis.
Purpose of the Study:
- To investigate if bacteriophage T4 morphogenesis can proceed without concurrent phage DNA replication.
- To analyze the impact of different media conditions on protein synthesis and phage assembly during inhibited DNA replication.
Main Methods:
- Infection of E. coli B with T4DtsLB3 under chloramphenicol treatment to block late protein synthesis.
- Inhibition of phage DNA replication using hydroxyurea and elevated temperature.
- Culturing in M9 or M9 with amino acids media to assess protein synthesis and assembly.
- Analysis of capsid protein composition using sucrose density gradients and chloroform lysis.
Main Results:
- In M9 medium, primarily early proteins were synthesized.
- In amino acid-rich medium, DNA synthesis was suppressed and degraded, but both early and late proteins were synthesized.
- Assembly of phage tails and capsids occurred, with some complete phage particles observed.
- Isolated capsids lacked specific proteins (Pwac, P23, P22) but contained others (Palt, P20, P23, P24).
- Viable phage yield ranged from 0.05% to 15% per cell.
Conclusions:
- Bacteriophage T4 morphogenesis is achievable without ongoing phage DNA replication.
- The presence of amino acids supports late protein synthesis and partial phage assembly even when DNA replication is blocked.