Immunoelectron microscopic localization of puromycin binding on the large subunit of the Escherichia coli ribosome.
Ribosomes from Escherichia coli strain Q13 have been photoaffinity labeled with [3H]puromycin in the presence of tetracycline. Puromycin-modified 50 S subunits appear to be identical with untreated subunits in electron micrographs and are precipitated by antibodies to the N6,N6'dimethyladenosine moiety of puromycin. Electron micrographs of subunit-antibody complexes show ribosomal subunits to which an individual antibody molecule is bound and pairs of subunits linked by an IgG molecule. Two regions of puromycin binding have been identified. The primary area, seen in 76% of the ribosome monomer complexes and 93% of the antibody-linked dimers, is beside (or on) the small central protuberance and on the side opposite the L7/L12 arm. A secondary area, maximally distant from the central protuberance, is seen in 22% of the monomeric complexes but only 7% of the antibody-linked dimers. In conjunction with our earlier localization of puromycin binding on the 30 S subunit (Olson, H. M., Grant, P. G., Glitz, D. G., and Cooperman, B. S. (1980) Proc. Natl. Acad. Sci. U. S. A. 77, 890-894), we now define a puromycin-binding neighborhood of the 70 S ribosome. In addition to providing evidence for the localization of the peptidyl transferase center within the 50 S subunit, our results contribute to the formulation of a model for tRNA binding to both 30 S subunits and 70 S ribosomes.
D000906:Antibodies; D000936:Antigen-Antibody Complex; D004926:Escherichia coli / Q000378:metabolism* / Q000648:ultrastructure; D008854:Microscopy, Electron; D008961:Models, Structural; D011691:Puromycin / Q000378:metabolism*; D012343:RNA, Transfer / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism* / Q000648:ultrastructure
D000906:Antibodies; D000936:Antigen-Antibody Complex; D011691:Puromycin; D012343:RNA, Transfer