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Image Search Results
Journal: The Journal of Biological Chemistry
Article Title: Proximity-dependent biotinylation detects associations between SARS coronavirus nonstructural protein 1 and stress granule-associated proteins
doi: 10.1016/j.jbc.2021.101399
Figure Lengend Snippet: Proximity-dependent labeling and pull-down identify the association between mutant nsp1 and cellular proteins. (A) BioID2-fused nsp1 proteins carrying mutations in the middle region of nsp1 were expressed, treated with 100 μM biotin for 16 hours and biotinylated proteins were isolated with streptavidin-coated magnetic bead. These proteins were separated in SDS gel followed by the analysis of specific protein association using immunoblot. (B) The above experiment was repeated with nsp1 carrying mutations in the structurally flexible C-terminus of the protein followed by immunoblot. C-terminus mutations weaken association by CPSF1, NUDT21, eIF4G.
Article Snippet: Antibodies were purchased from
Techniques: Labeling, Mutagenesis, Isolation, SDS-Gel, Western Blot
Journal: eLife
Article Title: An mRNA-binding channel in the ES6S region of the translation 48S-PIC promotes RNA unwinding and scanning
doi: 10.7554/eLife.48246
Figure Lengend Snippet: ( a ) Patterns of protein crosslinking generated with the indicated mRNAs, which are labeled with 4-thio-U and [α- 32 P]-GTP. The identified protein bands are indicated, with a relative quantification of the intensity of the protein bands shown below, according to this experiment and the one shown in . ( b ) Mapping of the interaction of eIF4A and eIF3g with mRNA in the ES6S region. SFV-DLP mRNAs with a single 4-thio-U placed at the indicated positions were used to assemble the 48S-PIC, and the pattern of protein crosslinking was analyzed. ( c ) The positions of mRNA that crosslinked with eIF4A and eIF3g were projected on the solvent side of the PIC, assuming that the mRNA is threaded along the ES6S region. The +11 position of mRNA is placed at the mRNA entry channel ( ; ). A rise per base of 4.5 Å for a stretched RNA strand was used as described previously .
Article Snippet: Western blots were carried out as described previously ( ) using the following primary antibodies: anti-RPS4X (sc-85133, Santa Cruz Biotech.), anti-RPS7 (sc-377317, Santa Cruz Biotech.), anti-RPS6 (sc-74576, Santa Cruz Biotech.), anti-eIF4A (STJ2724, St. John´s lab,),
Techniques: Generated, Labeling
Journal: eLife
Article Title: An mRNA-binding channel in the ES6S region of the translation 48S-PIC promotes RNA unwinding and scanning
doi: 10.7554/eLife.48246
Figure Lengend Snippet: ( a ) Denaturing immunoprecipitation (dIP) of crosslinked 48S-PIC revealed the presence of eIF4A and eIF3g bound to the indicated mRNAs (arrowheads). ( b ) Western blot analysis showing the specificity of the eIF4A1 antibody. HeLa cells were transfected with the indicated esiRNA, and 72 hr later, the resulting cells extracts were probed with anti-eIF4A1 and anti-eIF2 antibodies. esiRNA against the human ZC3HAV1 gene was used as control.
Article Snippet: Western blots were carried out as described previously ( ) using the following primary antibodies: anti-RPS4X (sc-85133, Santa Cruz Biotech.), anti-RPS7 (sc-377317, Santa Cruz Biotech.), anti-RPS6 (sc-74576, Santa Cruz Biotech.), anti-eIF4A (STJ2724, St. John´s lab,),
Techniques: Immunoprecipitation, Western Blot, Transfection, esiRNA
Journal: eLife
Article Title: An mRNA-binding channel in the ES6S region of the translation 48S-PIC promotes RNA unwinding and scanning
doi: 10.7554/eLife.48246
Figure Lengend Snippet: The amount of [ 32 P]-mRNA bound to WRF is indicated (upper panel), showing the pattern of protein crosslinking with the indicated mRNAs in the absence and presence of VIC–oligo 4. The unresolved eIF3g and eIF4A bands are shown (lower panel).
Article Snippet: Western blots were carried out as described previously ( ) using the following primary antibodies: anti-RPS4X (sc-85133, Santa Cruz Biotech.), anti-RPS7 (sc-377317, Santa Cruz Biotech.), anti-RPS6 (sc-74576, Santa Cruz Biotech.), anti-eIF4A (STJ2724, St. John´s lab,),
Techniques: