Figure 2

Hyaluronidase purification from the venom of V. dubius . (A) Gel filtration chromatography on Superdex® G-75 showing three active fractions. (B) Positive fractions from the previous step were applied to an affinity column of heparin-Sepharose (5 mL). The enzyme was detected at peak 2 (arrow), not retained by column. (C) C-18 reversed phase chromatography for final purification, using ACN gradient. (D) Left: SDS-PAGE of crude venom (Vd) and purified Hyase (Hy), using polyacrylamide gel 12% stained with silver nitrate. The estimated mass for the enzyme was 43 kDa. Right: crude venom and purified enzyme zymography. M = molecular weight markers.