Molecular cloning and characterization of the genes (pbpA and rodA) responsible for the rod shape of Escherichia coli K-12: analysis of gene expression with transposon Tn5 mutagenesis and protein synthesis directed by constructed plasmids
| Autor: | Michio Matsuhashi, Takahisa Ohta, Hiroshi Matsuzawa, S Asoh |
|---|---|
| Rok vydání: | 1983 |
| Předmět: |
DNA
Bacterial Transposable element Mutagenesis (molecular biology technique) Muramoylpentapeptide Carboxypeptidase Molecular cloning Biology Microbiology Plasmid Bacterial Proteins Escherichia coli Penicillin-Binding Proteins Cloning Molecular Molecular Biology Gene Genetics Regulation of gene expression Structural gene DNA Restriction Enzymes Chromosomes Bacterial Lambda phage biology.organism_classification Molecular biology Gene Expression Regulation Genes Hexosyltransferases Genes Bacterial Mutation Peptidyl Transferases DNA Transposable Elements Chromosome Deletion Carrier Proteins Plasmids Research Article |
| Zdroj: | Journal of Bacteriology. 154:10-16 |
| ISSN: | 1098-5530 0021-9193 |
| DOI: | 10.1128/jb.154.1.10-16.1983 |
| Popis: | Two cell shape-determining genes of Escherichia coli K-12, pbpA, the structural gene for penicillin-binding protein 2, and rodA, whose protein is unknown, were subcloned into plasmid vectors from the transducing phage lambda MAd lip24, which carries the lip-leuS region of the E. coli chromosome. Plasmids with restriction enzyme-created deletions or transposon Tn5 insertions were isolated, and studies of genetic complementation of these plasmids with chromosomal mutations were carried out. Thus, a physical and genetic map of the rodA-pbpA region was established. The genes rodA and pbpA lie side by side within a 4.4-kilobase-pair region. The size of the rodA gene has been shown to be between 0.86 and 1.6 kilobase pairs; such DNA would encode a protein with a molecular weight between 32,000 and 59,000. Since Tn5 mutagenesis of the rodA gene did not affect the expression of the pbpA gene and vice versa, the genes rodA and pbpA seem to have independent promoters. Analysis of the proteins synthesized from the constructed plasmids in maxicells revealed that the plasmid carrying the pbpA gene encoded penicillin-binding protein 2 and amplification of the protein occurred. The product of the rodA gene was not identified. |
| Databáze: | OpenAIRE |
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