Zorbax gf 250

The molecular mass of GusA, GusA-CBD, and CBD-GusA were examined by SDS-PAGE under denaturing conditions, using a pre-stained ladder (Bio-Rad) as a reference proteins. All protein bands were stained with Coomassie blue for visualization. The molecular mass of the native enzymes were estimated using an HPLC system (Agilent 1260, Santa Clara, CA) equipped with a UV detector at 210 nm using a ZORBAX GF-250 (250 mm × 4.6 mm, Agilent). The column was initially eluted with 130 mM NaCl/20 mM Na₂HPO₄ (pH 7.0) as the mobile phase. The flow rate was 1.0 ml/min and the column temperature was 23°C. The column was calibrated with thyroglobulin (669 kDa), aldolase (158 kDa), albumin (67 kDa), ovalbumin (43 kDa) and chymotrypsinogen A (25 kDa), as reference proteins, and the molecular mass of the native enzyme was calculated by comparing with the migration length of reference proteins. The molecular mass of the aggregated native enzymes were estimated after 12 h using a Superose_12 10/300 column for gel filtration chromatography (Amersham Biosciences). The enzyme solution was applied to the column and eluted with 50 mM Tris-HCl (pH 7.5) buffer containing 150 mM NaCl at a flow rate of 1 ml/min.