A specific role for the phosphorylation of mammalian acidic ribosomal protein P2.
The acidic ribosomal proteins P1-P2 from rat liver were overproduced for the first time by expression of their cDNA in Escherichia coli. They were tested for their ability to reactivate inactive P1-P2-deficient core particles derived from 60 S ribosomal subunits treated with dimethylmaleic anhydride, in poly(U)-directed poly(Phe) synthesis. The recombinant P1-P2 were unable to reactivate these core particles although they could bind to them. When recombinant P1-P2 had been phosphorylated first with casein kinase II, they were as efficient in the reactivation process as P1-P2 extracted with ethanol/KCl from the 60 S subunits. Reconstitution experiments were carried out using all possible combinations of the two recombinant proteins phosphorylated or not. Reactivation of the core particles required the presence of both P1 and P2 with the latter in its phosphorylated form. These experiments reveal a distinct role for P1 and P2 in protein synthesis. Phosphorylated P2 produced a partial quenching of the intrinsic fluorescence of eukaryotic elongation factor 2, which was not observed with the unphosphorylated protein. This result demonstrates the existence of an interaction between phosphorylated P2 and eukaryotic elongation factor 2. P2 also quenched part of the intrinsic fluorescence of P1, due to the interaction between the two proteins.
Vard
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Laboratoire de Biochimie Médicale, Institut de Biologie et Chimie des Protéines, Centre National de la Recherche Scientifique, 7, passage du Vercors, 69367 Lyon Cedex 07, France.
Guillot
D
D
Bargis
P
P
Lavergne
J P
JP
Reboud
J P
JP
D000818:Animals; D004926:Escherichia coli; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism; D005453:Fluorescence; D008099:Liver / Q000378:metabolism; D008299:Maleic Anhydrides / Q000378:metabolism; D008970:Molecular Weight; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D012269:Ribosomal Proteins / Q000378:metabolism*
D015852:Eukaryotic Initiation Factor-2; D008299:Maleic Anhydrides; D010750:Phosphoproteins; D012269:Ribosomal Proteins; C031937:phosphoprotein P2, ribosomal; C035690:ribosomal phosphoprotein P1; C007474:2,3-dimethylmaleic anhydride