Fig. 2.

Typical chromatograms of the purification procedure of latent and active cgAUS. Enzymatic assays for active cgAUS were performed by monitoring the oxidation of 1 ml 50 µm fisetin in 125 mm sodium citrate pH 5.5 at 280 nm (Jimenez et al. 1998). 2.5 mm SDS were added to the reaction mixture for latent and proteolytically cleaved latent cgAUS assays. One unit of enzyme was defined as the amount that catalyzed the formation of 1 nmol of oxidized fisetin per minute. a CEX chromatography using SP-Sepharose FF, pH 5.0. b AEX chromatography using Mono Q, pH 8.5. Fractions containing latent, proteolytically cleaved latent and active cgAUS are indicated below the chromatogram. c Polishing CEX chromatography of active sample 1 using Mono S, pH 5.0. d Polishing CEX chromatography of active samples 2 and 3 using Mono S, pH 5.0. e Polishing CEX chromatography of active sample 4 using Mono S, pH 5.0. f Polishing CEX chromatography of latent sample 5 using Mono S, pH 5.6