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Penetration of colicin M into cells of Escherichia coli
Abstract:
A new class of colicin M-tolerant mutants of Escherichia coli K-12 was isolated. The mutants exhibited an unusually high tolerance in that they were unaffected by colicin titers of 10(6). The tolerance was confined to colicin M. It was mapped at a locus called tolM, which is close to rpsL. The following gene order was determined: aroE, tolM, rpsL, cysG. The tolerance could be caused by a defect in the uptake of colicin M or by a mutation at the site of action. Insensitive tonA and tonB mutants became sensitive to colicin M upon treatment by osmotic shock, whereas the tolM mutants remained insensitive. Trypsin rescue experiments showed that the tonB-dependent uptake of colicin M required energy like the other tonB-related transport processes. When bound to energy-depleted cells, colicin M prevented adsorption of phage T5. The receptor became accessible to the phage when the cells were energized, except in tonB mutants. These data suggest that the function controlled by the tonB gene is required for the translocation of colicin M from its initial binding site at the tonA-coded receptor protein to the target.
Insights
New Escherichia coli K-12 mutants show high tolerance to colicin M, a potent bacterial toxin. This colicin M tolerance is linked to the tolM gene, impacting toxin uptake and action.
Area of Science:
- Microbiology
- Bacteriology
- Genetics
Background:
- Colicins are bacteriocins produced by Escherichia coli that inhibit the growth of other bacteria.
- Colicin M is a particularly potent colicin that inhibits protein synthesis.
- Understanding colicin resistance mechanisms is crucial for developing new therapeutic strategies.
Purpose of the Study:
- To isolate and characterize new mutants of Escherichia coli K-12 exhibiting tolerance to colicin M.
- To map the genetic locus responsible for colicin M tolerance.
- To elucidate the mechanism of colicin M uptake and action.
Main Methods:
- Isolation and characterization of colicin M-tolerant mutants.
- Genetic mapping of the tolM locus.
- Osmotic shock experiments.
- Trypsin rescue experiments.
- Phage adsorption assays.
Main Results:
- A new class of colicin M-tolerant mutants was isolated, unaffected by high colicin titers.
- The tolerance was mapped to the tolM locus, located near rpsL with the gene order aroE, tolM, rpsL, cysG.
- tolM mutants remained insensitive to colicin M after osmotic shock, unlike tonA and tonB mutants.
- Colicin M uptake is tonB-dependent and requires energy, similar to other tonB-related transport processes.
- Colicin M binding to the tonA receptor prevents phage T5 adsorption, which is reversed upon cell energization, except in tonB mutants.
Conclusions:
- The tolM gene is involved in colicin M tolerance.
- Colicin M uptake requires the function of the tonB gene for translocation from the tonA receptor to its target.
- This study provides insights into the mechanism of colicin M resistance and uptake in Escherichia coli.