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Published on: March 25, 2020
Entry exclusion in the IncHI1 plasmid R27 is mediated by EexA and EexB
James E Gunton1, John E R Ussher, Michelle M Rooker
1Department of Medical Microbiology and Immunology, 1-63 Medical Sciences Building, University of Alberta, Edmonton, Alberta, Canada T6G 2R3.
Plasmid
|January 15, 2008
Summary
Entry exclusion in IncH plasmids prevents redundant DNA transfer. Two genes, eexA and eexB, encode proteins that localize to bacterial membranes and mediate this plasmid incompatibility.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Conjugative plasmids utilize entry exclusion mechanisms to prevent redundant DNA transfer into recipient cells containing similar plasmids.
- The incompatibility H group (IncH) plasmid R27 is known to exhibit this exclusion phenomenon.
Purpose of the Study:
- To identify the genetic basis of entry exclusion in the IncH plasmid R27.
- To characterize the function and localization of the proteins involved in this mechanism.
Main Methods:
- Cosmid library construction and transformation of the R27 plasmid.
- Conjugation assays to screen for entry exclusion phenotype.
- Reverse-transcriptase analysis to identify genes within the Z operon.
- Sub-cloning and screening of individual genes.
- Bacterial fractionation to determine protein localization.
Main Results:
- The Z operon within the R27 plasmid's Tra2 region mediates entry exclusion.
- Two genes, eexA and eexB, were identified as independently responsible for inhibiting IncH-related plasmid entry.
- EexA protein is localized to the cytoplasmic membrane, and EexB protein to the outer membrane.
- Mutations in eexA and eexB abolished the exclusion phenotype, indicating their crucial role.
Conclusions:
- The IncH entry exclusion mechanism in R27 is mediated by the EexA and EexB proteins.
- These proteins likely interact to prevent redundant plasmid transfer.
- Understanding this mechanism provides insight into plasmid biology and evolution.
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