Figure 1.
Identification of elements within the N-terminus of Sro7 required for binding to the exocyst subunit Exo84. (A) Upper panel: Purified full length Sro7 and a N-terminal deletion of Sro7(aa 50–1033) were bound to immobilized GST-Exo84(aa 1–326) or GST alone in a pull-down assay. Coomassie of the GST-Exo84 fusion protein and quantitation of binding are shown on the right. Error bars represent SD; P values were obtained using a two-tailed Student’s t test with significance annotated as *, P < 0.05; **, P < 0.01; *** P < 0.001; ns = no significant difference. Lower panel: Purified full-length Sro7 and an N-terminal deletion of Sro7(aa 50–1033) were bound to GST-Sec9 or GST alone in a pull-down assay. Coomassie of the GST-Sec9 fusion protein and quantitation of binding are shown to the right. Error bars represent SD; P values were obtained as above. (B) Alignment of the N-terminus of Sro7 and Sro77 shows conserved amino acids between the yeast paralogs. Amino acids 1 through 10 are numbered. His-tagged wild-type Sro7-NT(aa 1–90) peptide and mutant Sro7-NT-A5,A6; Sro7-NT-A7,A8; Sro7-NT-A9,A10 and Sro7-NT-A(4-8) peptides were purified and then bound to immobilized GST-Exo84(aa 1–326) in a pull-down assay. Quantitation of binding is shown to the right. Error bars represent SD and P values were obtained as described above. (C) Purified His-tagged Exo84(aa 1–326) was bound to C-terminally tagged GST-Sro7-NT(aa 1–79) immobilized on beads using mutant GST-Sro7-NT-A5, A6 as a control. The additional bands in the binding sample for GST-Sro7-NT-A5,A6 were present on beads prior to binding. (D) Purified wild-type exocyst was bound to GST-Sro7-NT(aa 1–79) and mutant GST-Sro7-NT-A5,A6 immobilized on beads. Quantitation of binding for four subunits, two from each sub-complex is shown to the right. Error bars represent SD. Source data are available for this figure: SourceData F1.

Identification of elements within the N-terminus of Sro7 required for binding to the exocyst subunit Exo84. (A) Upper panel: Purified full length Sro7 and a N-terminal deletion of Sro7(aa 50–1033) were bound to immobilized GST-Exo84(aa 1–326) or GST alone in a pull-down assay. Coomassie of the GST-Exo84 fusion protein and quantitation of binding are shown on the right. Error bars represent SD; P values were obtained using a two-tailed Student’s t test with significance annotated as *, P < 0.05; **, P < 0.01; *** P < 0.001; ns = no significant difference. Lower panel: Purified full-length Sro7 and an N-terminal deletion of Sro7(aa 50–1033) were bound to GST-Sec9 or GST alone in a pull-down assay. Coomassie of the GST-Sec9 fusion protein and quantitation of binding are shown to the right. Error bars represent SD; P values were obtained as above. (B) Alignment of the N-terminus of Sro7 and Sro77 shows conserved amino acids between the yeast paralogs. Amino acids 1 through 10 are numbered. His-tagged wild-type Sro7-NT(aa 1–90) peptide and mutant Sro7-NT-A5,A6; Sro7-NT-A7,A8; Sro7-NT-A9,A10 and Sro7-NT-A(4-8) peptides were purified and then bound to immobilized GST-Exo84(aa 1–326) in a pull-down assay. Quantitation of binding is shown to the right. Error bars represent SD and P values were obtained as described above. (C) Purified His-tagged Exo84(aa 1–326) was bound to C-terminally tagged GST-Sro7-NT(aa 1–79) immobilized on beads using mutant GST-Sro7-NT-A5, A6 as a control. The additional bands in the binding sample for GST-Sro7-NT-A5,A6 were present on beads prior to binding. (D) Purified wild-type exocyst was bound to GST-Sro7-NT(aa 1–79) and mutant GST-Sro7-NT-A5,A6 immobilized on beads. Quantitation of binding for four subunits, two from each sub-complex is shown to the right. Error bars represent SD. Source data are available for this figure: SourceData F1.

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