It has been shown in vitro that Saccharomyces cerevisiae strand exchange protein 1 (Sep1) promotes the transfer of one strand of a linear duplex DNA to a homologous single-stranded DNA circle. Sep1 also has an exonuclease active on DNA and RNA. By using exonuclease III-treated linear duplex DNA with various lengths of single-stranded tail as well as Ca2+ to inhibit the exonuclease activity of Sep1, we show that the processivity of exonuclease activity of Sep1 is greater than previously reported. The results in this work also demonstrate that the joint molecule between the linear duplex and single-stranded circle observed from the Sep1-promoted strand transfer reaction is just the pairing between the long single-stranded tail of the linear duplex DNA (generated by the exonuclease activity of Sep1) and the single-stranded circular DNA. When a synthetic Holliday junction was used as substrate, branch migration facilitated by Sep1 could not be detected. Finally, using electron microscopy no alpha-structure, a joint molecule with displaced single-stranded DNA tail that indicates branch migration could be observed. The results imply that Sep1 cannot promote branch migration in vitro. Further investigation is needed to determine the role of Sep1 in recombination in vivo.

• PMID: 9478940
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Zhang Z, Simons AM, Prabhu VP, Chen J

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