Monoclonal antibodies against acidic phosphoproteins P0, P1, and P2 of eukaryotic ribosomes as functional probes.
Mice were immunized against ribosomal acidic proteins P1 and P2 from Artemia salina, and three kinds of monoclonal antibodies were isolated. One recognized P0 in addition to both P1 and P2 (anti-P). The other two recognized either P1 (anti-P1) or P2 (anti-P2) specifically and did not recognize P0. The anti-P antibody, but not anti-P1 or anti-P2, recognized a 22-amino acid peptide corresponding to the carboxyl-terminal sequence common to P1 and P2. This antibody, but not the others, inhibited poly(U)-directed polyphenylalanine synthesis. The anti-P1 bound to ribosomes but failed to inhibit polyphenylalanine synthesis: the anti-P2 did not bind to ribosomes at all. The anti-P and its Fab fragments inhibited the elongation step of protein synthesis, namely, the binding of elongation factors 1 alpha and 2 to ribosomes as well as their ribosome-coupled GTPase activities. Anti-P had little effect on the nonenzymatic phenylalanyl-tRNA binding to ribosomes and on peptidyltransferase activity. These results suggest the functional importance of the homologous carboxyl-terminal region of the three P proteins for the interaction of the ribosome with the two elongation factors. The epitope of anti-P1 must reside in a region of the protein which is not directly involved in its function.
Department of Biochemistry, Niigata University School of Medicine, Japan.
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Uchiumi
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Kominami
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000494:pharmacology; D000918:Antibody Specificity; D001156:Artemia / Q000032:analysis; D002499:Centrifugation, Density Gradient; D000939:Epitopes / Q000276:immunology; D005260:Female; D020963:GTP Phosphohydrolase-Linked Elongation Factors / Q000378:metabolism; D051379:Mice; D008814:Mice, Inbred NZB; D010452:Peptide Biosynthesis; D020648:Peptide Elongation Factor 1; D020652:Peptide Elongation Factor 2; D010445:Peptide Elongation Factors / Q000378:metabolism; D010455:Peptides; D010750:Phosphoproteins / Q000276:immunology*; D012269:Ribosomal Proteins / Q000276:immunology; D012270:Ribosomes / Q000276:immunology / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D020648:Peptide Elongation Factor 1; D020652:Peptide Elongation Factor 2; D010445:Peptide Elongation Factors; D010455:Peptides; D010750:Phosphoproteins; D012269:Ribosomal Proteins; C031937:phosphoprotein P2, ribosomal; C035690:ribosomal phosphoprotein P1; C054675:ribosomal protein P0; C012001:polyphenylalanine; D020963:GTP Phosphohydrolase-Linked Elongation Factors