Summary
It had been shown earlier, that RNA polymerase 13 S particles contain the large components with a molecular weight of about 3–105 and small subunits with a molecular weight of 4·104-1·105. These polymerase components easily dissociate and reassociate with restoration of the enzyme activity.
Both temperature-sensitive (tsX) and rifamycin-resistant (rif-r-I) mutations proved to affect the large polymerase component without changing the small subunits. These mutations were mapped at different, though closely linked, loci of metB-thi region of E. coli K12 chromosome. These results as well as certain literature data allow to conclude that the large RNA polymerase component consists of at least two polypeptides, one being altered by ts mutation, and the other—by rif-r mutation.
The large polymerase component when separated from the small subunits retain the ability to bind to T2 phage DNA while the separate small subunits lack this property. Rifamycin does not affect RNA polymerase-T2 DNA binding while ts mutation leads to inability of the enzyme to form stable complexes with DNA. Therefore, it is likely that the polypeptide affected by ts mutation is responsible for the attachment of RNA polymerase to specific sites of DNA template. On the other hand, the small subunits as well as polypeptide of the large component, which determines RNA polymerase sensitivity to rifamycin, seem not to participate in the enzyme binding to DNA template. It is suggested, that the catalytic site of RNA polymerase is located in the large component and formed by rifamycin-binding polypeptide. The small subunits are supposed to have regulatory function and activate the large components.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Babinet, C., Condamine, H.: Mutants résistants à la rifampicine, modifiés dans leur DNA-RNA-polymerase. C. R. Acad. Sci. (Paris) 267, 231–232 (1968).
Bautz, E. K. F., Dunn, J. J.: DNA-dependent RNA polymerase from phage T4 infected E. coli: an enzyme missing a factor required for transcription of T4 DNA. Biochem. Biphys. Res. Commun. 34, 230–237 (1969).
Berg, D., Barrett, K., Hinkle, D., McGrath, J., Chamberlin, M.: A subunit of RNA polymerase involved in chain initiation. Fed. Proc. 28, 659 (1969).
Bogdanova, E. S., Zograf, Yu. N., Bass, I. A., Goldfarb, A. D.: RNA polymerase from uninfected and phage infected cells of E. coli. Symp. “Structure and genetic functions of biopolymers”, Publ. by Kurchatov Institute of Atomic Energy, Moscow 1969.
Bremer, H., Konrad, M., Bruner, R.: Capacity of T4 DNA to serve as template for purified Escherichia coli RNA polymerase. J. molec. Biol. 16, 104–117 (1966).
Burgess, R. R., Travers, A. A., Dunn, J. J., Bautz, E. K. F.: Factor stimulating transcription by RNA polymerase. Nature (Lond.) 221, 43–46 (1969).
Cohen, S. N., Maitra, U., Hurwitz, J.: Role of DNA in RNA synthesis. XI. Selective transcription of λ DNA segments in vitro by RNA polymerase of Escherichia coli. J. molec. Biol. 26, 19–38 (1967).
Colvill, A. J., Kanner, L. C., Tocchini-Valentini, G. P., Sarnat, M. T., Geiduschek, E. P.: Asymmetric RNA synthesis in vitro: heterologus DNA-enzyme systems. Proc. nat. Acad. Sci. (Wash.) 53, 1140 (1965).
Crawford, L. V., Crawford, E. M., Richardson, J. P., Slayter, H. S.: The binding of RNA polymerase to polyoma and papilloma DNA. J. molec. Biol. 14, 593–597 (1965).
Ezekiel, D. H., Hutchins, J. E.: Mutations affecting RNA polymerase associated with rifampicin resistance in Escherichia coli. Nature (Lond.) 220, 276–277 (1968).
Geiduschek, E. P., Snyder, L., Colvill,A. J., Sarnat, M.: Selective synthesis of T-even bacteriophage early messenger in vitro. J. molec. Biol. 19, 541 (1966).
Igarshi, K., Yura, T.: The role of RNA polymerase in genetic transcription. Biochem. biophys. Res. Commun. 34, 65–69 (1969).
Ishihama, A., Hurwitz, J.: Alteration and reversible dissociation of E. coli RNA polymerase. Fed. Proc. 28, 659 (1969).
Jones, O. W., Berg, P.: Studies on the binding of RNA polymerase to polynucleotides. J. molec. Biol. 22, 199–209 (1966).
Kamsolova, S. G., Manyakov, V. F., Kisselev, N. A., Shemyakin, M. F., Astaurova, O. B., Khesin, R. B.: Studies on complexes between RNA polymerase and DNA. Molec. Biologia 3, 74–87 (1969).
Khesin, R. B., Mindlin, S. Z., Gorlenko, Zh. M., Ilyina, T. S.: Temperature sensitive mutations affecting RNA synthesis in Escherichia coli. Molec. Gen. Genetics 103, 194–208 (1968).
—, Shemyakin, M. F., Gorlenko, Zh. M., Bogdanova, S. L., Afanasieva, T. P.: RNA polymerase in E. coli cell infected with T2 phage. Biochimiya 27, 1092–1105 (1962).
— Mindlin, S. Z., Ilyina, T. S.: Studies on the RNA polymerase of Escherichia coli K12 with use of mutation affecting its activity. Molec. Biologia 2, 898–907 (1968).
—: Studies on the RNA polymerase in Escherichia coli K12 using the mutation affecting its activity. J. molec. Biol. 42, 401–411 (1969).
Lennox, C. S.: Transduction of linked genetic characters of the host by bacteriophage P1. Virology 1, 190–206 (1955).
Mauro, E. di, Snyder, L., Marino, P., Lamberti, A., Coppo, A., Tocchini-Valentini, G. P.: Rifampicin sensitivity of the components of DNA-dependent RNA polymerase. Nature (Lond.) 222, 533–537 (1969).
Neuhoff, V., Schill, W. B., Sternbach, H.: Mikro-disk-elektrophoretische Analyse reiner DNA-abhängiger RNA-Polymerase aus Escherichia coli, III. Differenzierung zwischen Syntheseort und Bindungsort für die Matrize. Hoppe-Seylers Z. physiol. Chem. 350, 335–340 (1969).
Richardson, J. P.: The binding of RNA polymerase to DNA. J. molec. Biol. 21, 83–112 (1966).
Satoshi, M., Hisaji, W., Kazuo, N., Hamao, U.: Inhibition of DNA-dependent RNA polymerase reaction of Escherichia coli by an antimicrobial antibiotic, streptovaricin. Biochim. biophys. Acta (Amst.), 157, 322–332 (1968).
Shemyakin, M. F., Manyakov, V. F., Kisselev, N. A., Khesin, R. B.: Synthesis of RNA on both strands of depolymerized DNA. Biochimiya 31, 594–604 (1966).
Sippel, A., Hartmann, G.: Mode of action of rifamycin on the RNA polymerase reaction. Biochim. biophys. Acta (Amst.) 157, 218–219 (1968).
Snyder, L., Geiduschek, E. P.: In vitro synthesis of T4 late messenger RNA. Proc. nat. Acad. Sci. (Wash.) 59, 459–466 (1968).
Tocchini-Valentini, G. P., Marino, P., Colvill, A. J.: A mutant of E. coli containing an altered DNA-dependent RNA polymerase. Nature (Lond.) 220, 275–276 (1968).
Travers, A. A., Burgess, R. R.: Cyclic re-use of the RNA polymerase sigma factor. Nature (Lond.) 222, 537–540 (1969).
Wehrli, W., Knüsel, F., Schmid, K., Staehelin, M.: Interaction of rifamycin with bacterial RNA polymerase. Proc. nat. Acad. Sci. (Wash.) 61, 667–673 (1968).
Wu, C. W.: Initiation by the RNA polymerase. Fed. Proc. 28, 659 (1969).
Yura, T., Igarashi, K.: RNA polymerase mutants of Escherichia coli, I. Mutants resistant to streptovaricin. Proc. nat. Acad. Sci. (Wash.) 61, 1313–1319 (1968).
Author information
Authors and Affiliations
Additional information
Communicated by D. Goldfarb
Rights and permissions
About this article
Cite this article
Khesin, R.B., Gorlenko, Z.M., Shemyakin, M.F. et al. Studies on the functions of the RNA polymerase components by means of mutations. Molec. Gen. Genetics 105, 243–261 (1969). https://doi.org/10.1007/BF00337475
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00337475