Fig. 5.

Binding and UV crosslinking of Bzm-ShhN in the presence of heparin. (A) Binding of modified ShhN proteins (50 μg) to a heparin-affinity column and elution with a linear gradient of 0 – 2.0 M NaCl in 5 mM sodium phosphate pH 5.5, 150 mM NaCl, 0.5 mM DTT buffer. (B) Bzm-ShhN was incubated with heparin and subjected to UV light (30 min, dark, 365 nm). Controls included ShhN + heparin, Bzm-ShhN alone or no UV. Crosslinking, SDS-PAGE and Western blotting carried out as described in Methods. Western blot with anti-ShhN polyclonal Ab. Cross-linked bands are indicated by vertical bar.