FIG. 3.

Purification of the CcaR protein from S. clavuligerus(pB17B) by HiTrap heparin-agarose. (A) The ammonium sulfate (40 to 60%) precipitate obtained from the cell extract was suspended in 10 mM phosphate buffer, pH 7.0, and 10.5 ml (105 mg of protein) of the suspension was applied to a heparin-agarose column. The proteins were eluted with a 0-to-2 M NaCl gradient. Solid line, protein; dashed line, NaCl. (Inset) Fractions giving positive signals in immunoblotting assays with anti-rCcaR antibodies. (B) Gel mobility shift of the ccaR-360 DNA fragment containing the ccaR promoter with fractions eluted from the heparin-agarose column. (C) Mobility shift of the DI-782 DNA fragment containing the cefD-cmcI bidirectional promoter with the same fractions eluted from the heparin-agarose column.