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Identification and biosynthesis of the bacteriophage T4 mot regulatory protein
Abstract:
The T4 mot gene regulates middle mode RNA synthesis in phage-infected cells. The mot gene product has been identified in two ways. (i) Infections with amber and temperature-sensitive mot mutants both lead to the disappearance of a number of protein bands on SDS-polyacrylamide gels. These are middle mode proteins whose synthesis depends on mot function. The mot protein disappears from such gels after infection with a mot amber mutant, but not with the mot missense mutant. (ii) This same protein is the only one to have a charge alteration when proteins from wild-type phage and mot missense mutant infections are compared by two-dimensional gel electrophoresis. Mot protein is basic and has a mol. wt. of 24 000. It migrates between the positions of gp 1 and gp IPIII on 15% SDS-polyacrylamide gels. Mot protein synthesis begins immediately after infection and continues until 4 min after infection at 30 degrees C, after which time it is strongly inhibited. This inhibition depends neither on T4 DNA synthesis nor on ADP ribosylation of the alpha subunits of the Escherichia coli RNA polymerase. The mot protein does not regulate its own biosynthesis. It is stable throughout the course of infection.
Insights
The T4 mot gene product regulates middle mode RNA synthesis in phage-infected cells. This essential regulatory protein was identified using mutant analysis and advanced gel electrophoresis techniques.
Area of Science:
- Molecular Biology
- Virology
- Genetics
Background:
- The T4 bacteriophage employs intricate regulatory mechanisms to control gene expression during infection.
- Middle mode RNA synthesis is a critical phase in the phage life cycle, requiring specific regulatory proteins.
Purpose of the Study:
- To identify and characterize the T4 mot gene product, responsible for regulating middle mode RNA synthesis.
- To elucidate the properties and synthesis regulation of the mot protein.
Main Methods:
- Utilizing amber and temperature-sensitive mot mutants to analyze protein expression changes via SDS-polyacrylamide gel electrophoresis.
- Employing two-dimensional gel electrophoresis to compare protein profiles between wild-type and mot missense mutant infections.
- Determining the molecular weight and charge characteristics of the mot protein.
Main Results:
- The mot gene product was identified as a basic protein with a molecular weight of 24,000 Da.
- Mutant analysis revealed that mot protein synthesis is essential for middle mode protein production.
- Two-dimensional gel electrophoresis identified a charge alteration in the mot protein from missense mutants.
- Mot protein synthesis is initiated early post-infection and is subsequently inhibited, independent of T4 DNA synthesis or RNA polymerase modification.
Conclusions:
- The T4 mot protein is a key regulator of middle mode RNA synthesis.
- The mot protein's synthesis is tightly regulated and does not auto-regulate its own production.
- The identified mot protein is stable throughout the infection cycle.