Binding of Met-tRNA-f to native and derived 40S ribosomal subunits.
Our previous work has shown that the native 40S ribosomal subunits (those found free in the cell sap) but not polyribosomal 40S subunits have additional associated proteins that are removed by 0.5 M KCl. In this communication we present evidence that in the Ehrlich cell one of the native subunit associated proteins is the mammalian initiation factor that forms a Met-tRNA-f-factor-GTP complex, and is required for the binding of Met-tRNA-f to the 40S subunit. Initial examination of the KCl wash of the Ehrlich cell total ribosomal pellet revealed a factor which (1) shifted the elution of Met-tRNA-f and of GTP from the included to the excluded volume on Sephadex G-100 chromatography, (2) stimulated the binding of Met-tRNA-f to Millipore filters, and (3) stimulated the binding of Met-tRNA-f to salt-washed 40S subunits. These activities were dependent upon or enhanced by GTP; were inhibited by GDP; were much greater for Met-tRNA-f than for Met-tRNA-m or for lysyl-tRNA; and were concentrated in the KCl ribosomal wash and were not detected in the cell soluble fraction. Met-tRNA-f bound in conjunction with a specific amount of KCl wash protein, to form a distinctive particle of bouyant density 1.40 g cm minus 3 in CsCl, identical in density to one form of the native 40S subunit. Native 40S subunits, but no other subunits, contained a factor which was eluted by 0.5 M KCl and which (1) stimulated the binding of Met-tRNA-f to Millipore filters, and (2) stimulated the binding of Met-tRNA-f to salt-washed 40S subunits. The factor appeared to be localized on the native 40S subunit of density 1.40 g cm minus 3.
D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism; D000818:Animals; D002286:Carcinoma, Ehrlich Tumor / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D004926:Escherichia coli / Q000201:enzymology; D008099:Liver / Q000378:metabolism; D008239:Lysine; D008715:Methionine; D051379:Mice; D011189:Potassium Chloride; D012343:RNA, Transfer / Q000378:metabolism; D051381:Rats; D012270:Ribosomes / Q000378:metabolism*; D014462:Ultrafiltration
D011189:Potassium Chloride; D012343:RNA, Transfer; D008715:Methionine; D000604:Amino Acyl-tRNA Synthetases; D008239:Lysine