Journal: bioRxiv
Article Title: Human Pannexin 1 Channel is NOT Phosphorylated by Src Tyrosine Kinase at Tyr199 and Tyr309
doi: 10.1101/2023.09.10.557063
Figure Lengend Snippet: A , In vitro phosphorylation of hPANX1-GFP WT by active human Src-GST protein. The leftmost panel showed the in-gel fluorescence of GFP overlaid with protein marker. The other panels (from left to right) represent the western blot signal of anti-PANX1, anti-pY100, anti-Src, anti-PANX1-PANX1-pY198, and anti-PANX1-pY308 antibodies, respectively. B , In vitro phosphorylation of hPANX1-GFP WT, L306A, K307A, V308A, Y309F, E310A, I311A, or L312A mutants. The left panel showed the in-gel fluorescence of GFP overlaid with protein marker. The right panel showed the western blot signal of anti-PANX1-pY308 antibody. Only hPANX1 WT lane showed western blot signal. C , In vitro phosphorylation of hPANX1-GFP WT, Y309A, or Y309F mutants. The left panel showed the in-gel fluorescence of GFP overlaid with protein marker (red). The right panel showed the western blot signal of anti-PANX1-pY308 antibody. The Y309A mutant rendered PANX1 prone to oligomerization in our SDS gel experiment, but not for Y309F. D , Whole cell lysate of HEK293T cells transiently transfected with hPANX1 WT or Y309F. Left panel showed the in-gel fluorescence of GFP overlaid with protein marker. Middle and right panels represent western blot result using anti-PANX1 and anti-PANX1-pY308 antibody, respectively. The location where PANX1 is expected to migrate in the SDS-PAGE gel is indicated. Of note, visualizing such a band requires a very long exposure, which resulted in a large number of non-specific bands. Figure 3—source data 1: Uncropped immunoblots for . Figure 3—source data 2: Raw image for in-gel fluorescence (GFP and mCherry) in . Figure 3—source data 3: Raw image for anti-PANX1 immunoblot in . Figure 3—source data 4: Raw image for anti-pY100 immunoblot in . Figure 3—source data 5: Raw image for anti-Src immunoblot in . Figure 3—source data 6: Raw image for anti-PANX1-pY198 immunoblot in . Figure 3—source data 7: Raw image for anti-PANX1-pY308 immunoblot in . Figure 3—source data 8: Raw image for in-gel fluorescence (GFP and mCherry) in . Figure 3—source data 9: Raw image for anti-PANX1-pY308 immunoblot in . Figure 3—source data 10: Raw image for in-gel fluorescence (GFP and mCherry) in . Figure 3—source data 11: Raw image for anti-PANX1 immunoblot in . Figure 3—source data 12: Raw image for anti-PANX1-pY308 immunoblot in .
Article Snippet: The in vitro phosphorylation is conducted using the human Src-GST protein (Sigma-Aldrich, Catalogue number: S1076) according to the manufacturer’s protocol.
Techniques: In Vitro, Fluorescence, Marker, Western Blot, Mutagenesis, SDS-Gel, Transfection, SDS Page
Sino Biological is a verified supplier 
Sino Biological manufactures this product 
