The selective release of one of the two L7/L12 dimers from the Escherichia coli ribosome induced by a monoclonal antibody to the NH2-terminal region.
Two monoclonal antibodies against different epitopes in Escherichia coli ribosomal protein L7/L12 were prepared and characterized as reported previously (Sommer, A., Etchison, J.R., Gavino, G., Zecherle, N., Casiano, C., and Traud, R.R. (1985) J. Biol. Chem. 260, 6522-6527). Both antibodies strongly inhibited polyuridylic acid-directed polyphenylalanine synthesis, ribosome-dependent GTPase activity, and the binding of elongation factor G to the ribosome at mole ratios over ribosomes of 4:1 or less. One epitope was shown to be within residues 1-73 (Ab 1-73) and the other within 74-120 (Ab 74-120). Incubation of 50 S ribosomal subunits or 70 S ribosomes with Ab 1-73, but not with Ab 74-120, leads to a partial loss of L7/L12 from the particle with no loss of any other protein. The experiment was repeated with ribosomes reconstituted with pure radioactive L7/L12 of determined specific activity in order to quantify the L7/L12 in the antibody-treated particle. The protein-deficient core particles isolated by sucrose gradient centrifugation after incubation with Ab 1-73 were found to contain, on average, two copies of L7/L12 and one Ab 1-73. The constancy of this stoichiometry in many experiments and the demonstration of Ab 1-73 on all particles indicate the presence of a homogeneous population of ribosomes, each with only one of the two L7/L12 dimers originally present. The results show a difference in the interactions of the two dimers with the ribosome and present a means of preparing ribosomes with one dimer in a specific binding site. The accompanying paper (Olson, H.M., Sommer, A., Tewari, D. S., Traut, R.R., and Glitz, D.G. (1986) J. Biol. Chem. 261, 6924-6932) shows by immune electron microscopy the location of the two antibody-binding sites and the effect of Ab 1-73 on structure.
D S
DS
Tewari
A
A
Sommer
R R
RR
Traut
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D001426:Bacterial Proteins / Q000378:metabolism*; D000939:Epitopes / Q000032:analysis; D004926:Escherichia coli / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D012269:Ribosomal Proteins / Q000276:immunology / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*; D013462:Sulfur Radioisotopes
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D001426:Bacterial Proteins; D000939:Epitopes; D046911:Macromolecular Substances; D012269:Ribosomal Proteins; D013462:Sulfur Radioisotopes; C016700:ribosomal protein L7-L12