Fig. 3. Purification of rimaPCI. rimaPCI was expressed in E. coli and retrieved from the culture media. After a disulfide reshuffling procedure and a clarification step, rimaPCI was purified by a mixed-mode chromatography followed by a gel filtration chromatography. (A) Protein elution profile from Streamline Direct HST adsorbent. Fractions enriched in imaPCI (gray area limited by dotted line in the chromatogram) were pooled and repurified by chromatography. (B) Gel filtration polishing of the mixed-mode chromatography eluate using a HiLoad 26/60 Superdex 30 prep grade column. The collected fraction of rimaPCI is indicated in the chromatogram by the gray area. (C) Tricine- SDS PAGE analysis of purified rimaPCI (7.5 µg); rPCI (7.5 µg) was also loaded in the gel for comparison purposes. Proteins were stained with Coomassie Brillant Blue. (D) RPHPLC elution profiles of purified rimaPCI (bold line) and rPCI (dotted line).