| Products/Services Used | Details | Operation |
|---|---|---|
| Gene Synthesis | with N-terminal His6 tags were made by inserting DNA between NcoI and BamHI sites of pET15b (GenScript, Piscataway, NJ). pET15b-His6-Nsp1(SARS CoV-2) was then used by GenScript to generate the substitution mutants described in the text./The vector pET15b-His6-eIF3g for expression in E. coli of human eIF3g (Genbank Acc. No. NM_003755) with an N-terminal His6 tag was made by inserting DNA between NdeI and XhoI sites of pET15b (GenScript). pET15b-His6-eIF3g was used by GenScript to generate the deletion and substitution mutants. | Get A Quote |
SARS CoV-2 nonstructural protein 1 (Nsp1) is the major pathogenesis factor that inhibits host translation using a dual strategy of impairing initiation and inducing endonucleolytic cleavage of cellular mRNAs. To investigate the mechanism of cleavage, we reconstituted it in vitro on β-globin, EMCV IRES and CrPV IRES mRNAs that use unrelated initiation mechanisms. In all instances, cleavage required Nsp1 and only canonical translational components (40S subunits and initiation factors), arguing against involvement of a putative cellular RNA endonuclease. Requirements for initiation factors differed for these mRNAs, reflecting their requirements for ribosomal attachment. Cleavage of CrPV IRES mRNA was supported by... More
