Fig. 8. Hg2+-mediated chemical crosslinking of BbZIP to validate the proposed OFC.
a Crosslinking of the A95C/L217C variant. The protein was purified in DDM, incubated with HgCl2 and then applied to non-reducing SDS-PAGE. The shifted bands after Hg treatment are indicated by arrows. The same batch of the purified protein was treated differently as indicated to confirm that the band shift is attributed to cysteine reacting with Hg2+. The single cysteine variants (A95C and L217C) were treated in the same way as the double variant. b Crosslinking of the double cysteine variants (V167C/V272C, W168C/V272C, and L169C/V272C) with mutations on the opposite side of residues 95 and 217. Note that the W168C/V272C variant could not be crosslinked by HgCl2 under the same condition. The residues subjected to mutagenesis are mapped to the IFC and the OFC model, respectively. The Cβ atoms of the residues are depicted in sphere mode. Source data are provided as a Source Data file.