Homologous Recombination

Microbial Genetics

Functions

• Double-Strand Break (DSB) repair

• Gene exchange (genetic diversity)

• Adaptation and evolution

Requirements

• Homologous DNAs must align

• Recombination synapse forms (DNA pairing complex)

• Heteroduplex DNA forms (strand exchange region)

• Rec proteins required

• ATP energy needed

Single Cross-Over Recombination

Results in integration of F plasmid into the bacterial chromosome → forms Hfr strain (high-frequency recombination)

Holliday Model

• Nicks occur in each DNA molecule

• Double-strand invasion occurs

• Heteroduplex region forms

• Holliday junction (cross-shaped intermediate) forms

• Branch migration extends heteroduplex

• Isomerization changes junction structure

• Resolution of the Holliday junction produces reciprocal exchange of alleles

Single-Strand Invasion (D-loop) Model

• Random nick in DNA 1

• Single-stranded end from DNA 1 invades DNA 2

• Synapse forms between the two DNAs

• Heteroduplex and Holliday junction follow similar steps as the Holliday model

RecB

slow helicase (top strand)

• ~100 copies bind to the ssDNA end

• Forms RecA–ssDNA filament

• Searches for homologous region in DNA 2

• Creates triple-strand complex in the major groove

• Promotes synapse formation and heteroduplex formation

RecD

fast helicase (bottom strand)

RecC

nuclease

Chi site (χ site)

halts nuclease activity

DSB Repair

Produces free single-stranded DNA end that initiates recombination

RuvA/B

recognize and bind the Holliday junction

Branch migration

driven by RuvB helicase

RuvC

resolves the Holliday junction, completing recombination