The alpha and gamma subunits of initiation factor eIF-2 can be cross-linked to 18S ribosomal RNA within the quaternary initiation complex, eIF-2.Met-tRNAf.GDPCP.small ribosomal subunit.
Initiation factor eIF-2 from rat liver was reacted with the hetero-bifunctional cross-linking reagents ABAI or APTPI without diminishing its ability to form the quaternary initiation complex with Met-tRNAf, GDPCP and the small ribosomal subunit. Upon irradiation with UV light, subunits alpha and gamma of eIF-2 became covalently linked to 18S ribosomal RNA. The subunits were identified electrophoretically after isolation of the covalent protein-rRNA complexes and subsequent degradation of the rRNA by nuclease and alkali treatments. The close proximity of the two factor subunits to sequences of ribosomal RNA within the quaternary complex could be confirmed in a second set of experiments using unmodified, 125I-labeled factor and diepoxybutane as cross-linking reagent.
D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D003432:Cross-Linking Reagents; D015852:Eukaryotic Initiation Factor-2; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000378:metabolism / Q000528:radiation effects; D007096:Imidoesters; D008715:Methionine; D010442:Peptide Chain Initiation, Translational; D011506:Proteins / Q000378:metabolism* / Q000528:radiation effects; D012335:RNA, Ribosomal / Q000378:metabolism*; D012343:RNA, Transfer / Q000378:metabolism* / Q000528:radiation effects; D012346:RNA, Transfer, Amino Acyl; D051381:Rats; D012270:Ribosomes / Q000378:metabolism*; D014466:Ultraviolet Rays
D003432:Cross-Linking Reagents; D015852:Eukaryotic Initiation Factor-2; D007096:Imidoesters; D011506:Proteins; D012335:RNA, Ribosomal; D012346:RNA, Transfer, Amino Acyl; C025207:eIF-2 Met-tRNA(f)(met) GDPCP; C027906:methyl-4-azidobenzoylaminoacetimidate; C027907:methyl-5-(4-azidophenyl)-4,5-dithiapentanimidate; D006160:Guanosine Triphosphate; D012343:RNA, Transfer; D008715:Methionine