No sliding during homology search by RecA protein

Abstract

The RecA protein of Escherichia coli is a prototype of the RecA/Rad51 family of proteins that exist in virtually all the organisms. In a process called DNA synapsis, RecA first polymerizes onto a single-stranded DNA (ssDNA) molecule; the resulting RecA-ssDNA complex then searches for and binds to a double-stranded DNA (dsDNA) molecule containing the almost identical, or "homologous, " sequence. The RecA-ssDNA complex thus can be envisioned as a sequence-specific binding entity. How does the complex search for its target buried within nonspecific sequences? One possible mechanism is the sliding mechanism, in which the complex first binds to a dsDNA molecule nonspecifically and then linearly diffuses, or slides, along the dsDNA. To understand the mechanism of homology search by RecA, this sliding model was tested. A plasmid containing four homologous targets in tandem was constructed and used as the dsDNA substrate in the synapsis reaction. If the sliding is the predominant search mode, the two outermost targets should act as more efficient targets than the inner targets. No such positional preference was observed, indicating that a long range sliding of the RecA-ssDNA complex does not occur. These and other available data can be adequately explained by a simple three-dimensional random collision mechanism.