Alteration of χ recognition by RecBCD reveals a regulated molecular latch and suggests a channel-bypass mechanism for biological control

Liang Yang, Naofumi Handa, Bian Liu, Mark S. Dillingham, Dale B. Wigley, Stephen C. Kowalczykowski

Research output: Contribution to journalArticlepeer-review

21 Scopus citations


The RecBCD enzyme is a complex heterotrimeric helicase/nuclease that initiates recombination at double-stranded DNA breaks. In Escherichia coli, its activities are regulated by the octameric recombination hotspot, χ (5′-GCTGGTGG), which is read as a singlestranded DNA sequence while the enzyme is unwinding DNA at over ∼1,000 bp/s. Previous studies implicated the RecC subunit as the "χ-scanning element" in this process. Site-directed mutagenesis and phenotypic analyses identified residues in RecC responsible for χ recognition [Handa N, et al., (2012) Proc Natl Acad Sci USA, 10.1073/pnas.1206076109]. The genetic analyses revealed two classes of mutants. Here we use ensemble and single-molecule criteria to biochemically establish that one class of mutants (type 1) has lost the capacity to recognize χ (lost-recognition), whereas the second class (type 2) has a lowered specificity for recognition (relaxed-specificity). The relaxed-specificity mutants still recognize canonical χ, but they have gained the capacity to precociously recognize single-nucleotide variants of χ. Based on the RecBCD structure, these mutant classes define an α-helix responsible for χ recognition that is allosterically coupled to a structural latch. When opened, we propose that the latch permits access to an alternative exit channel for the single-stranded DNA downstream of χ, thereby avoiding degradation by the nuclease domain. These findings provide a unique perspective into the mechanism by which recognition of a single-stranded DNA sequence switches the translocating RecBCD from a destructive nuclease to a constructive component of recombinational DNA repair.

Original languageEnglish (US)
Pages (from-to)8907-8912
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number23
StatePublished - Jun 5 2012


  • Allosteric switch
  • Protein-DNA interactions

ASJC Scopus subject areas

  • General


Dive into the research topics of 'Alteration of χ recognition by RecBCD reveals a regulated molecular latch and suggests a channel-bypass mechanism for biological control'. Together they form a unique fingerprint.

Cite this