Related Experiment Videos
Quantification of group A colicin import sites
1Laboratorie d'Ingénierie et de Dynamique des Systèmes Membranaires, Centre National de la Recherche Scientifique, UPR 9027, Marseille, France.
Journal of Bacteriology
|September 1, 1995
Summary
A pore-forming colicin A mutant, unable to form channels, still binds receptors. This finding helps estimate the number of colicin A receptors and translocation sites on Escherichia coli cells.
Area of Science:
- Microbiology
- Molecular Biology
- Biophysics
Background:
- Pore-forming colicins are bacteriocins that create ion channels in the Escherichia coli inner membrane.
- Colicin translocation involves outer membrane receptors and translocation machinery, with two groups (A and B) utilizing different protein pathways (Tol or TonB/ExbB/ExbD).
Purpose of the Study:
- To investigate the role of the pore-forming domain in colicin translocation and receptor interaction.
- To quantify the number of colicin A receptors and translocation sites on Escherichia coli.
Main Methods:
- Utilized a double-cysteine colicin A mutant with a disulfide bond in its pore-forming domain.
- Measured colicin-induced K+ efflux and performed competition experiments with wild-type colicins and the mutant.
Main Results:
- The colicin A mutant was translocated through the envelope but could not form an inner membrane channel.
- Preincubation with the mutant blocked group A colicin binding but not group B, indicating specific interactions.
- The mutant remained associated with the receptor and import machinery during inner membrane interaction.
Conclusions:
- Escherichia coli cells possess approximately 400 colicin A receptors.
- Each Escherichia coli cell has approximately 1,000 colicin A translocation sites.