Summary
The location of the ribosomal proteins L14, L17, L18, L19, L22 and L23 on the surface of the 50S subunit of E. coli ribosomes was determined by immune electron microscopy. Antibodies (bivalent IgG's) specific for the six ribosomal proteins were used to form 50S subunit dimers (50S-IgG-50S). The dimers were separated from non-bound antibodies and 50S subunit-monomers and larger aggregates by sucrose density gradient centrifugation. The attachment of each of the six immunoglobulins to 50S subunits was visualized directly by electron microscopy using negative staining and correlated with one or more of the structural features of the particle. Each of the proteins was found to occupy a unique position. Proteins L14, L19 and L23 are located in the region of the 50S subunit which interacts with the 30S particle. Proteins L17 and L22 are on the opposite side of the 50S ribosomal subunit, whilst L18 holds a position on one of the lateral protuberances of a crown-like shaped 50S subunit.
Two main forms of 50S subunits were seen on the electron micrographs: one of them had a crown-like shape; the second revealed kidney shaped images. It was demonstrated that these two images are only projections of one structure of the subunit that resembled an armchair; each of the forms could be transformed into the other through rotation by 90°.
The results allowed the proposal of a three-dimensional model of the 50S subunit with the location of six different ribosomal proteins as illustrated in Fig. 13.
Similar content being viewed by others
References
Benjamini, E., Michaeli, D., Young, J. D.: Antigenic determinants of proteins of defined sequences. In: Current topics in microbiology and immunology, vol. 58. Berlin-Heidelberg-New York: Springer 1972
Finch, J. T., Klug, A.: The structure of viruses of the Papilloma-Polyoma type. III. Structure of rabbit Papilloma virus. Topography of contrast in negative-staining for electron-microscopy. J. molec. Biol. 13, 1–12 (1965)
Garrett, R. A., Müller, S., Spierer, P., Zimmermann, R. A.: Binding of 50S ribosomal subunit proteins to 23S RNA of E. coli. J. molec. Biol. 88, 553–557 (1974)
Garrett, R. A., Wittmann, H. G.: Structure of bacterial ribosomes Advanc. Protein Chem. 27, 277–347 (1973)
Gesteland, R. F.: Isolation and characterisation of ribonuclease I mutants of E. coli. J. molec. Biol. 16, 67–84 (1966)
Gray, P. N., Bellemare, G., Monier, R., Garrett, R. A., Stöffler, G.: Identification of the nucleotide sequences involved in the interaction between Escherichia coli 5S RNA and specific 50S subunit proteins. J. molec. Biol. 77, 133–152 (1973)
Gray, P. N., Garrett, R. A., Stöffler, G., Monier, R.: An attempt at the identification of the proteins involved in the incorporation of 5S RNA during 50-S ribosomal subunit assembly. Europ. J. Biochem. 28, 412–421 (1972)
Hart, R. G.: Electron microscopy of the 50S ribosomes of Escherichia coli. Biochim. biophys. Acta (Amst.) 60, 629–637 (1962)
Highland, J. H., Ochsner, E., Gordon, J., Bodley, J., Hasenbank, R., Stöffler, G.: Inhibition of elongation factor G function by antibodies specific for several ribosomal proteins. Proc. nat. Acad. Sci. (Wash.) 71, 627–630 (1974)
Hindennach, I., Kaltschmidt, E., Wittmann, H. G.: Ribosomal proteins. Isolation of proteins from 50S ribosomal subunits of Escherichia coli. Europ. J. Biochem. 23, 12–16 (1971a)
Hindennach, I., Stöffler, G., Wittmann, H. G.: Ribosomal proteins. Isolation of the proteins from 30S ribosomal subunits of Escherichia coli. Europ. J. Biochem. 23, 7–11 (1971b)
Horne, R. J., Erdmann, V.: ATPase and GTPase activities associated with a specific 5S RNA-protein complex. Proc. nat. Acad. Sci. (Wash.) 70, 2870–2873 (1973)
Huxley, H. E., Zubay, G.: Electron microscope observations on the structure of microsomal particles from Escherichia coli. J. molec. Biol. 2, 10–18 (1960)
Kurland, C. G.: Structure and function of the bacterial ribosome. Ann. Rev. Biochem. 41, 377–408 (1972)
Lubin, M.: Observations on the shape of the 50S ribosomal subunit. Proc. nat. Acad. Sci. (Wash.) 61, 1454–1461 (1968)
Maschler, R.: Immunochemische Untersuchungen zur Funktion der Einzelproteine der 30S-Untereinheit von Escherichia coli-Ribosomen. Inaugural-Dissertation, Freie Universität, Berlin (1973)
Morrison, C. A., Garrett, R. A., Zeichhardt, H., Stöffler, G.: Proteins occurring at, or near, the subunit interface of E. coli ribosomes. Molec. gen. Genet. 127, 359–368 (1973)
Nirenberg, M. W., Matthaei, J. H.: The dependence of cell-free protein synthesis in E. coli upon naturally occurring or synthetic polyribonucleotides. Proc. nat. Acad. Sci. (Wash.) 47, 1588–1602 (1961)
Pilz, I., Kratky, O., Licht, A. Sela, M.: Shape and volume of anti-poly (d-alanyl) antibodies in the presence and absence of tetra-d-alanine as followed by small-angle X-ray scattering. Biochemistry (Wash.) 12, 4998–5005 (1973)
Pongs, O., Nierhaus, K. H., Erdmann, V. A., Wittmann, H. G.: Active sites in Escherichia coli ribosomes. FEBS Letters 40, S28-S37 (1974)
Sarma, V. R., Silverton, E. W., Davis, D. R., Terry, W. D.: The three-dimensional structure at 6 Å resolution of a human immune-globulin molecule. J. biol. Chem. 246, 3753–3759 (1971)
Stöffler, G.: Structure and function of the Escherichia coli ribosome. The use of immuno-chemical methods for the analysis. In: The ribosome (P. Lengyel, M. Nomura A. Tissières, eds.). New York: Cold Spring Harbor in press 1974
Stöffler, G., Hasenbank, R., Lütgehaus, M., Maschler, R., Morrison, C. A., Zeichhardt, H., Garrett, R. A.: The accessibility of proteins of the Escherichia coli 30S ribosomal subunit to antibody binding. Molec. gen. Genet. 127, 89–110 (1973)
Stöffler, G., Wittmann, H. G.: Sequence differences of Escherichia coli 30S ribosomal proteins as determined by immunochemical methods. Proc. nat. Acad. Sci. (Wash.) 69, 2283–2287 (1971a)
Stöffler, G., Wittmann, H. G.: Ribosomal proteins, XXV. Immunological studies on Escherichia coli ribosomal proteins. J. molec. Biol. 62, 407–409 (1971b)
Tischendorf, G. W., Zeichhardt, H., Stöffler, G.: Location of proteins S5, S13 and S14 on the surface of the 30S ribosomal subunit from Escherichia coli as determined by immune electron microscopy. Molec. gen. Genet. 134, 209–223 (1974)
Wabl, M. R.: Electron microscopic localization of two proteins on the surface of the 50S ribosomal subunit of Escherichia coli using specific antibody markers. J. molec. Biol. 84, 241–247 (1974)
Wabl, M. R., Barends, P. J., Nanninga, N.: Tilting experiments with negatively stained E. coli ribosomal subunits. An electron microscopic study. Cytobiologie 7, 1–9 (1973b)
Wabl, M. R., Doberer, H. G., Höglund, S., Ljung, L.: Electron microscopic study on isolated 30S ribosomal subunits of Escherichia coli. Cytobiologie 7, 111–115 (1973a)
Weber, H. J.: Stoichiometric measurements of 30S and 50S ribosomal proteins from Escherichia coli. Molec. gen. Genet., 119, 233–248 (1972)
Wittmann, H. G., Stöffler, G.: Structure and function of bacterial ribosomal proteins. In: Protein biosynthesis (L. Bosch, ed.), Amsterdam: North Holland Publ. Co. 1972
Wittmann, H. G., Wittmann-Liebold, B.: Chemical structure of bacterial ribosomal proteins. In: The ribosome (P. Lengyel, M. Nomura, A. Tissières, eds.). New York: Gold Spring Harbor, in press 1974
Author information
Authors and Affiliations
Additional information
Communicated by E. Bautz
Rights and permissions
About this article
Cite this article
Tischendorf, G.W., Zeichhardt, H. & Stöffler, G. Determination of the location of proteins L14, L17, L18, L19, L22 and L23 on the surface of the 50S ribosomal subunit of Escherichia coli by immune electron microscopy. Molec. Gen. Genet. 134, 187–208 (1974). https://doi.org/10.1007/BF00267715
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00267715