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. 2009 Jun 16;4(6):e5935. doi: 10.1371/journal.pone.0005935

Figure 3. SNX4 interacts with CHC via a clathrin box variant.

Figure 3

(a) Schematic view of SNX4 wt, SNX4 198-451Δ and SNX4 109-113Δ. (b) Cells transfected as indicated were lysed and proceeded to anti-myc immunoprecipitation for 4 h at 4°C. The immunoprecipitate and whole cell lysate (WCL) were separated by SDS-PAGE and analyzed by Western blot with the indicated antibodies. Quantification of CHC binding corrected for expression level of the myc constructs is shown to the right, n = 3. (c) The amino acid sequences of GST-peptides are shown with putative clathrin box in red. The numbering represents amino acid number in the SNX4 sequence. GST pull-down was performed with cell lysate at 4°C overnight. The pull-down was subjected to SDS-PAGE and analyzed by Western blot with the indicated antibodies. GST alone was used as negative control. (d) Putative clathrin boxes of SNX1, SNX2, SNX3, SNX4 (105–116) and reverse clathrin box of Hrs are aligned. GST pull-down was performed as in (c) with the indicated peptides. (e) Schematic view of GST-peptides containing the putative clathrin box (red) and point mutations (blue). GST pull-down was performed as in (c) with the indicated peptides.