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Structural basis for initiation of transcription from an RNA polymerase–promoter complex

Nature volume 399, pages 80–83 (1999)Cite this article

A Correction to this article was published on 01 July 1999

Abstract

Although the single-polypeptide-chain RNA polymerase from bacteriophage T7 (T7RNAP), like other RNA polymerases, uses the same mechanism of polymerization as the DNA polymerases, it can also recognize a specific promoter sequence, initiate new RNA chains from a single nucleotide, abortively cycle the synthesis of short transcripts, be regulated by a transcription inhibitor, and terminate transcription1,2,3. As T7RNAP is homologous to the Pol I family of DNA polymerases4, the differences between the structure of T7RNAP complexed to substrates and that of the corresponding DNA polymerase complex provides a structural basis for understanding many of these functional differences. T7RNAP initiates RNA synthesis at promoter sequences that are conserved from positions −17 to +6 relative to the start site of transcription. The crystal structure at 2.4 Å resolution of T7RNAP complexed with a 17-base-pair promoter shows that the four base pairs closest to the catalytic active site have melted to form a transcription bubble. The T7 promoter sequence is recognized by interactions in the major groove between an antiparallel β-loop and bases. The amino-terminal domain is involved in promoter recognition and DNA melting. We have also used homology modelling of the priming and incoming nucleoside triphosphates from the T7 DNA-polymerase ternary complex structure to explain the specificity of T7RNAP for ribonucleotides, its ability to initiate from a single nucleotide, and the abortive cycling at the initiation of transcription.

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Figure 1: Surface representation of the T7RNAP–promoter complex structure.
Figure 2: Interactions between the T7-promoter DNA and three regions of T7RNAP that are unique to the RNA polymerase.
Figure 3: Homology-modelled initiation complex.

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Acknowledgements

We thank many members of the T.A.S. laboratory for assistance with data collection at beamlines X12C (NSLS, Brookhaven Laboratory), A1, F1 and F2 (Cornell High Energy Synchrotron). This work was supported by a grant from the N.I.H.

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Author notes

  1. David Jeruzalmi

    Present address: Laboratory of Molecular Biophysics, The Rockefeller University, 1230 York Avenue, New York, New York, 10021, USA

Authors and Affiliations

  1. Departments of Molecular Biophysics and Biochemistry, 266 Whitney Avenue, New Haven, 06520-8114, Connecticut, USA

    Graham M. T. Cheetham, David Jeruzalmi & Thomas A Steitz

  2. Departments of Chemistry, 266 Whitney Avenue, New Haven, 06520-8114, Connecticut, USA

    Thomas A Steitz

  3. Howard Hughes Medical Institute, Yale University, 266 Whitney Avenue, New Haven, 06520-8114, Connecticut, USA

    Graham M. T. Cheetham & Thomas A Steitz

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Cheetham, G., Jeruzalmi, D. & Steitz, T. Structural basis for initiation of transcription from an RNA polymerase–promoter complex. Nature 399, 80–83 (1999). https://doi.org/10.1038/19999

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