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The EMBO Journal (1998) 17, 4511–4526, doi:10.1093/emboj/17.15.4511 Figure 7 Protein domains and conformational changes in the activation of RepA, a DNA replication initiator Rafael Giraldo, José M. Andreu and Ramón Díaz-Orejas

Figure 7 EMSA of pPS10 ds-DNA sequences in complex with RepA (or its fragments). (A) A 39 bp (32P-labelled) double-stranded oligonucleotide, including the repA operator sequence (ds-1IR), was incubated at 510-9 M with increasing amounts (ramp headers, from 510-9 to 510-6 M) of either purified RepA–WT or each of its fragments, and then run in a 6% polyacrylamide native gel. The autoradiograph shows two complexes for the WT protein (CWT1 and CWT2) (see text), and none for any of the RepA fragments. (B) The same type of experiment was performed with 110-6 M of a radiolabelled 45 bp double-stranded oligonucleotide including a single iteron plus the DnaA box (ds-1DR). DNA was incubated with purified RepA–WT or each of its fragments (concentration range from 510-8 to 510-6 M). Low-affinity single protein–DNA complexes appear only for RepA–WT (CWT), N37 (well) and N132 (CN132).

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