Figure 1.

(a) Purification of CLBPah from N. naja atra venom. 0.5 g of dried crude venom was dissolved in 10 ml buffer A (0.02 M NaH₂PO₄/Na₂HPO₄ pH 6.0), centrifuged at 10 000 rev min⁻¹ to remove insoluble materials and then applied onto a CM-Sepharose column (1 × 20 cm) pre-equilibrated with the same buffer. The elution was performed at a flow rate of 0.5 ml min⁻¹ with a combined gradient of pH value and salt concentration produced by mixing 200 ml buffer A with 200 ml buffer B (0.02 M Na₂HPO₄/NaH₂PO₄ pH 8.0 containing 0.5 M NaCl) followed by two column volumes of buffer B. The ninth fraction, containing CLBPah, was pooled and desalted for further isolation. The absorbance at 280 nm is indicated by a solid line and the salt gradient by a dashed line. (b) Purification of CLBPah on a gel-filtration column. Fraction 9 was applied onto a Sephadex G-50 column (1.6 × 100 cm) and eluted with 0.15 M NaCl solution. Peak 3 is CLBPah. The inset shows SDS–PAGE analysis of CLBPah. Lane 1, CLBPah. Lane 2, molecular-weight markers: rabbit phosphorylase b (97 400 Da), bovine serum albumin (66 200 Da), rabbit actin (43 000 Da), bovine carbonic anhydrase (31 000 Da), trypsin inhibitor (20 100 Da) and hen egg-white lysozyme (14 400 Da).