Figure 4.

In vitro interaction of single missense mutant proteins as compared to the leucine zipper motif of chHAT-1 with the GST–chCAF-1p48 fusion protein. (A) A series of single missense mutant proteins, as compared to the leucine zipper motif of chHAT-1, were constructed and then their binding activities, as compared to the GST–chCAF-1p48 fusion protein, were examined. The mutant proteins used are denoted by appropriate abbreviations. (B) An in vitro immunoprecipitation experiment on the interaction of the single missense mutant proteins of chHAT-1 with the Flag-tagged chCAF-1p48 protein was carried out using [³⁵S]Met-labeled Flag-tagged chCAF-1p48 and [³⁵S]Met-labeled chHAT-1, or missense mutants of it, followed by the addition of anti-Flag M2 beads. Aliquots of eluates from the affinity beads were analyzed by 12% SDS–PAGE and the gels were then subjected to fluorography (b). A portion (20%) of each reaction mixture was removed before the immunoprecipitation as an input sample (a).