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KMID : 0578319920020020239
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Molecules and Cells
1992 Volume.2 No. 2 p.239 ~ p.243
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Effects of DNA Deletions and Insertions on Strand Exchange Reactions Mediated by E. coli RecA Protein
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Phue Je-Nie
Hahn Tae-Ryong
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Abstract
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RecA protein from E. coli promotes the homologous pairing and strand exchange between single-stranded circular and homologous linear duplex DNA. This strand exchange reaction generates nicked circular heteroduplex and linear single-stranded DNA. In consequence, the completion of strand exchange can be demonstrated by the production of nicked circular heteroduplex DNA which can be detected by gel electrophoresis. E. coli strain KM4104 which contains a multicopy plasmid carrying E. coli recA gene was used for the purification of RecA protein. Effects of deletions and insertions in DNA substrates on RecA mediated strand exchange reactions were investigated in vitro by the purified RecA protein. RecA catalyzed strand exchange to form stable heteroduplex products with upto 575 unpaired bases when the insert is located within a single stranded circular molecule. However, when the insert is located in the linear duplex DNA substrates, exchange products were formed with upto 72 unpaired bases but those were not detected with more than 120 base pairs of insertions. These results contrast with previous finding by others that RecA protein spans insertions with hundreds base pairs in the duplex DNA [Bianchi, M. E., and Radding, C. M. (1983) Cell 35, 511-5201.
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