Related Experiment Videos
Primary structure of colicin M, an inhibitor of murein biosynthesis
Abstract:
The DNA sequence of the colicin M activity gene cma was determined. A polypeptide consisting of 271 amino acids was deduced from the nucleotide sequence. The amino acid sequence agreed with the peptide sequences determined from the isolated colicin. The molecular weight of active colicin M was 29,453. The primary translation product was not processed. In the domain required for uptake into cells, colicin M contained the pentapeptide Glu-Thr-Leu-Thr-Val. A similar sequence was found in all colicins which are taken up by a TonB-dependent mechanism and in outer membrane receptor proteins which are constituents of TonB-dependent transport systems. The structure of colicin M in the carboxy-terminal activity domain had no resemblance to the pore-forming colicins or colicins with endonuclease activity. Instead, the activity domain contained a sequence which exhibited homology to the sequence around the serine residue in the active site of penicillin-binding proteins of Escherichia coli. The colicin M activity gene was regulated from an SOS box upstream of the adjacent colicin B activity gene on the natural plasmid pColBM-Cl139.
Insights
The colicin M activity gene (cma) DNA sequence reveals a 271-amino acid protein. Colicin M
Area of Science:
- Bacteriocin research
- Molecular biology
- Genetics
Background:
- Colicins are bacteriocins produced by Escherichia coli.
- Colicin M is a protein toxin that inhibits cell wall synthesis.
- The mechanism of colicin M uptake and activity is not fully understood.
Purpose of the Study:
- To determine the DNA sequence of the colicin M activity gene (cma).
- To deduce the amino acid sequence and analyze the structure-function relationship of colicin M.
- To investigate the regulation of the colicin M gene.
Main Methods:
- DNA sequencing of the colicin M gene (cma).
- Amino acid sequence analysis and comparison with known colicins and proteins.
- Identification of functional domains and regulatory elements.
Main Results:
- The cma gene encodes a 271-amino acid polypeptide with a molecular weight of 29,453 Da.
- Colicin M possesses a cell uptake domain with a conserved pentapeptide (Glu-Thr-Leu-Thr-Val), characteristic of TonB-dependent transport.
- The activity domain shows homology to penicillin-binding proteins, suggesting a novel mechanism of action distinct from pore-forming or endonuclease colicins.
- The cma gene is regulated by an SOS box, linked to the colicin B gene regulation on plasmid pColBM-Cl139.
Conclusions:
- The deduced amino acid sequence of colicin M is consistent with experimental data.
- Colicin M utilizes a TonB-dependent mechanism for cell entry.
- Colicin M's activity domain suggests a unique inhibitory mechanism targeting bacterial cell wall synthesis, possibly via transpeptidase inhibition.
- Gene regulation of colicin M is coordinated with colicin B through an SOS response.