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Figure 3 | Journal of Biomedical Science

Figure 3

From: The N-terminal domain of Escherichia coli RecA have multiple functions in promoting homologous recombination

Figure 3

Production of wild-type and mutant RecA proteins. An improved SUMO fusion protein expression system was used to produce authentic RecA protein. (A) Purified RecA protein on a 12% SDS-PAGE gel stained with Coomassie Blue. (B) Nuclease activity assay. Purified RecA protein (1 μM) or exonuclease I (20 units; New England Biolabs) was incubated with 5'-end 32P-labeled P1656 ssDNA (50 nucleotides, 3 μM), respectively. The reaction mixtures were treated with proteinase K and then electrophoresed on 20% native acrylamide gels. Gels were visualized on a phosphorimager, with overexposure to confirm that the purified RecA proteins exhibited no detectable nuclease activity. (C) Purified RecA proteins are capable of polymerization into nucleoprotein helical filaments. Negative-staining electron microscopy images show various RecA proteins with a ΦX174 dsDNA substrate in the presence of ATPγS. Scale bars (in black) are 100 nm.

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