Figure 4.

Pathways for integrative recombination. (A) Recombination reactions were performed by using attP DNA radiolabeled on both ends, mixing all DNA and protein components concurrently at the temperature indicated with no preincubation (attP + attB//+ mIHF + Int) and with incubation at the temperature indicated for 0.1 to 30 min. As a control, the intasome was preformed on ice by preincubation of attP, Int-L5 (as indicated), and mIHF, and attB was added later (as indicated) and incubated on ice for 30 min (attP + mIHF + Int//+ attB). At the 10- and 30-min room temperature time points, a doublet can be distinguished near the free attP position, which is the formation of the attL product complex after strand exchange (7). (B) Multiple assembly pathways. At the top are shown the reaction components attP, attB, Int, and mIHF. On the left is shown the synapsis late pathway in which Int and mIHF first form an intasome complex that then captures attB DNA to generate SC1. Further addition of mIHF promotes folding of this open structure into a postulated compact but active complex in which strand exchange occurs to release the products, free attR DNA, and an attL complex. On the right is shown the alternative synapsis early pathway, in which the initial complex formed is SC2, followed by assembly into complex 1 or perhaps directly into the closed active complex. The only one of these complexes not identified by native gel electrophoresis is the putative active closed complex, which is postulated to form slowly (such that complex 1 accumulates) but is rapidly converted into products.