Fig. 3. Impact of select mutations on the binding between TAC-1 and ZYG-9. (A) GST pull-down experiments with wild-type, M58I or L229F GST-TAC-1 and in vitro translated [³⁵S]-labeled wild-type, I862K or E1317K ZYG-9 TBR (654-1415), as indicated. Top panels show 10% of the input material and 100% of the corresponding pulled-down material detected by autoradiography. The bottom panels show the Coomassie Blue staining of the GST-TAC-1 fusions used in each case, indicating that comparable amounts of proteins had been added. The weaker band detected in the input lanes (denoted by 10%) corresponds to a 60-kDa protein from the in vitro transcription/translation mix. GST alone did not retain significant amounts of ZYG-9 TBR (data not shown). (B) Quantification of TAC-1-ZYG-9 binding, expressed as the percentage of each radioactive product retained on Glutathione-sepharose beads, averaged from three independent experiments, ± s.e.m. 100% corresponds to binding observed with wild-type ZYG-9 TBR and wild-type GST-TAC-1.