  Homologous recombination plays a key role in generating genetic diversity,
while maintaining protein functionality. The mechanisms by which RecA enables a
single-stranded segment of DNA to recognize a homologous tract within a whole
genome are poorly understood. The scale by which homology recognition takes
place is of a few tens of base pairs, after which the quest for homology is
over. To study the mechanism of homology recognition, RecA-promoted homologous
recombination between short DNA oligomers with different degrees of heterology
was studied in vitro, using fluorescence resonant energy transfer. RecA can
detect single mismatches at the initial stages of recombination, and the
efficiency of recombination is strongly dependent on the location and
distribution of mismatches. Mismatches near the 5' end of the incoming strand
have a minute effect, whereas mismatches near the 3' end hinder strand exchange
dramatically. There is a characteristic DNA length above which the sensitivity
to heterology decreases sharply. Experiments with competitor sequences with
varying degrees of homology yield information about the process of homology
search and synapse lifetime. The exquisite sensitivity to mismatches and the
directionality in the exchange process support a mechanism for homology
recognition that can be modeled as a kinetic proofreading cascade.
