%0 Journal Article
%T The RecB helicase-nuclease tether mediates Chi hotspot control of RecBCD enzyme
%A Gerald R Smith
%A Susan K Amundsen
%J Archive of "Nucleic Acids Research".
%D 2019
%R 10.1093/nar/gky1132
%X In bacteria, repair of DNA double-strand breaks uses a highly conserved helicase¨Cnuclease complex to unwind DNA from a broken end and cut it at specific DNA sequences called Chi. In Escherichia coli the RecBCD enzyme also loads the DNA strand-exchange protein RecA onto the newly formed end, resulting in a recombination hotspot at Chi. Chi hotspots regulate multiple RecBCD activities by altering RecBCD¡¯s conformation, which is proposed to include the swinging of the RecB nuclease domain on the 19-amino-acid tether connecting the helicase and nuclease domains. Here, we altered the tether and tested multiple RecBCD activities, genetically in cells and enzymatically in cell-free extracts. Randomizing the amino-acid sequence or lengthening it had little effect. However, shortening it by as little as two residues or making substitutions of ¡Ý10 proline or ¡Ý9 glycine residues dramatically lowered Chi-dependent activities. These results indicate that proper control of RecBCD by Chi requires that the tether be long enough and appropriately flexible. We discuss a model in which the swing-time of the nuclease domain determines the position of Chi-dependent and Chi-independent cuts and Chi hotspot activity
%U https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6326792/