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Preparation of the Mgm101 Recombination Protein by MBP-based Tagging Strategy
Published on: June 25, 2013
The RecBC enzyme loads RecA protein onto ssDNA asymmetrically and independently of chi, resulting in constitutive
J J Churchill1, D G Anderson, S C Kowalczykowski
1Biochemistry and Molecular Biology Graduate Group, University of California, Davis, California 95616-8665 USA.
Abstract:
Double-strand DNA break repair and homologous recombination in Escherichia coli proceed by the RecBCD pathway, which is regulated by cis-acting elements known as chi sites. A crucial feature of this regulation is the RecBCD enzyme-directed loading of RecA protein specifically onto the 3'-terminal, chi-containing DNA strand. Here we show that RecBC enzyme (lacking the RecD subunit) loads RecA protein constitutively onto the 3'-terminal DNA strand, with no requirement for chi. This strand is preferentially utilized in homologous pairing reactions. We propose that RecA protein loading is a latent property of the RecBCD holoenzyme, which is normally blocked by the RecD subunit and is revealed following interaction with chi.
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