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Published on: January 8, 2015
Isolation of the gene encoding the Hin recombinational enhancer binding protein
R C Johnson1, C A Ball, D Pfeffer
1Department of Biological Chemistry, School of Medicine, University of California, Los Angeles 90024.
Summary
The Fis protein is essential for efficient Hin-mediated DNA inversion in Escherichia coli. While not required for cell growth, Fis facilitates high rates of site-specific DNA inversion in vivo.
Area of Science:
- Molecular Biology
- Genetics
- Microbiology
Background:
- Hin-mediated DNA inversion in vitro requires a cis-acting recombinational enhancer and the Fis protein.
- The Fis protein binds to the recombinational enhancer sequence.
Purpose of the Study:
- To clone and determine the primary sequence of the gene encoding the Fis protein.
- To investigate the role of Fis in Hin-mediated site-specific DNA inversion in vivo.
Main Methods:
- Cloning and sequencing of the fis gene.
- Deduced amino acid sequence analysis to identify functional motifs.
- Construction of mutant Escherichia coli strains lacking a functional fis gene.
- Assessment of Hin-mediated site-specific inversion rates in wild-type and mutant strains.
Main Results:
- The fis gene was cloned, and its primary sequence was determined.
- The deduced Fis amino acid sequence (98 amino acids) revealed a potential helix-turn-helix DNA binding motif.
- The fis gene maps to 72 min on the Escherichia coli chromosome.
- Mutant strains lacking fis are viable but exhibit significantly reduced rates of Hin-mediated DNA inversion.
Conclusions:
- Fis is a DNA-binding protein with a helix-turn-helix motif, crucial for Hin-mediated DNA inversion.
- Fis is not essential for Escherichia coli viability under standard laboratory conditions.
- Fis plays a critical role in facilitating high-efficiency, site-specific DNA inversion in vivo.
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