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Association of the rIIA protein with the bacterial membrane
Abstract:
Cell membrane proteins synthesized after infection of Escherichia coli B with wild-type phage T4 and rIIA mutants were analyzed by dodecyl sulfate-polyacrylamide gel electrophoresis. A protein with an approximate molecular weight of 74,000 is present in membranes isolated from T4r(+)-infected cells, but is not found in membranes prepared from cells infected with an rIIA mutant in which the major part of the rIIA cistron is deleted. In addition, infection of E. coli B with different rIIA amber mutants and deletions gives peptides, which are associated with the bacterial membrane, of molecular weights consistent with the location of the respective mutations in the cistron. The rIIA protein is synthesized with delayed early kinetics. The synthesis of the rIIB protein, which is also located in the membrane, is not affected by mutations in the A cistron; conversely, synthesis of the rIIA protein is not affected by mutations in the B cistron. A mutant (rII 1589) contains a deletion that originates in the A cistron and extends into the adjacent B cistron. This mutant directs the synthesis of a compound membrane protein consisting of the undeleted portions of the A and B cistrons. The synthesis of the compound protein appears to be under the control of the A promoter.
Insights
Phage T4 infection of Escherichia coli reveals a 74,000 MW membrane protein linked to the rIIA gene. Mutations within the rIIA cistron produce specific peptides, indicating gene function in membrane protein synthesis.
Area of Science:
- Molecular Biology
- Virology
- Bacteriology
Background:
- Bacteriophage T4 infection alters host cell membrane protein synthesis.
- The rIIA and rIIB cistrons of phage T4 encode membrane proteins.
Purpose of the Study:
- To analyze cell membrane proteins synthesized after Escherichia coli B infection with wild-type phage T4 and rIIA mutants.
- To investigate the relationship between rIIA gene mutations and the synthesis of specific membrane proteins.
Main Methods:
- Dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was used to analyze cell membrane proteins.
- Infection of E. coli B with various phage T4 rIIA mutants and wild-type.
Main Results:
- A 74,000 MW protein was identified in membranes of T4r(+)-infected cells, absent in cells infected with a major rIIA deletion mutant.
- rIIA amber mutants and deletions produced peptides of molecular weights correlating with mutation locations.
- rIIA protein synthesis follows delayed early kinetics.
- rIIA and rIIB gene products are synthesized independently.
- A deletion mutant (rII 1589) synthesized a compound membrane protein from both rIIA and rIIB cistrons, controlled by the rIIA promoter.
Conclusions:
- The rIIA gene product is a 74,000 MW membrane protein.
- Specific mutations within the rIIA cistron lead to the production of distinct membrane-associated peptides.
- The rIIA and rIIB genes function independently in protein synthesis, but can produce a combined protein product when deletions occur.