A Mechanism of Nucleotide Misincorporation During Transcription Due to Template-strand Misalignment

Overview

Journal Mol Cell

Publisher Cell Press

Specialty Cell Biology

Date 2006 Oct 21

PMID 17052458

Citations 22

Authors

Richard T Pomerantz

Dmitry Temiakov

Michael Anikin

Dmitry G Vassylyev

William T McAllister

Affiliations

Soon will be listed here.

Abstract

Transcription errors by T7 RNA polymerase (RNAP) may occur as the result of a mechanism in which the template base two positions downstream of the 3' end of the RNA (the TSn+1 base) is utilized during two consecutive nucleotide-addition cycles. In the first cycle, misalignment of the template strand leads to incorporation of a nucleotide that is complementary to the TSn+1 base. In the second cycle, the template is realigned and the mismatched primer is efficiently extended, resulting in a substitution error. Proper organization of the transcription bubble is required for maintaining the correct register of the DNA template, as the presence of a complementary nontemplate strand opposite the TSn+1 base suppresses template misalignment. Our findings for T7 RNAP are in contrast to related DNA polymerases of the Pol I type, which fail to extend mismatches efficiently and generate predominantly deletion errors as a result of template-strand misalignment.

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References

Suzuki M, Avicola A, Hood L, Loeb L . Low fidelity mutants in the O-helix of Thermus aquaticus DNA polymerase I. J Biol Chem. 1997; 272(17):11228-35. DOI: 10.1074/jbc.272.17.11228. View

Bell J, ECKERT K, Joyce C, Kunkel T . Base miscoding and strand misalignment errors by mutator Klenow polymerases with amino acid substitutions at tyrosine 766 in the O helix of the fingers subdomain. J Biol Chem. 1997; 272(11):7345-51. DOI: 10.1074/jbc.272.11.7345. View

Kiefer J, Mao C, Braman J, Beese L . Visualizing DNA replication in a catalytically active Bacillus DNA polymerase crystal. Nature. 1998; 391(6664):304-7. DOI: 10.1038/34693. View

Remington K, Bennett S, Harris C, Harris T, Bebenek K . Highly mutagenic bypass synthesis by T7 RNA polymerase of site-specific benzo[a]pyrene diol epoxide-adducted template DNA. J Biol Chem. 1998; 273(21):13170-6. DOI: 10.1074/jbc.273.21.13170. View

Li Y, Kong Y, Korolev S, Waksman G . Crystal structures of the Klenow fragment of Thermus aquaticus DNA polymerase I complexed with deoxyribonucleoside triphosphates. Protein Sci. 1998; 7(5):1116-23. PMC: 2144016. DOI: 10.1002/pro.5560070505. View

Minnick D, Bebenek K, Osheroff W, Turner Jr R, Astatke M, Liu L . Side chains that influence fidelity at the polymerase active site of Escherichia coli DNA polymerase I (Klenow fragment). J Biol Chem. 1999; 274(5):3067-75. DOI: 10.1074/jbc.274.5.3067. View

Doublie S, Ellenberger T . The mechanism of action of T7 DNA polymerase. Curr Opin Struct Biol. 1999; 8(6):704-12. DOI: 10.1016/s0959-440x(98)80089-4. View

Salas M . Resolution of head-on collisions between the transcription machinery and bacteriophage phi29 DNA polymerase is dependent on RNA polymerase translocation. EMBO J. 1999; 18(20):5675-82. PMC: 1171634. DOI: 10.1093/emboj/18.20.5675. View

Westover K, Bushnell D, Kornberg R . Structural basis of transcription: nucleotide selection by rotation in the RNA polymerase II active center. Cell. 2004; 119(4):481-9. DOI: 10.1016/j.cell.2004.10.016. View

10.

Gong X, Zhang C, Feig M, Burton Z . Dynamic error correction and regulation of downstream bubble opening by human RNA polymerase II. Mol Cell. 2005; 18(4):461-70. DOI: 10.1016/j.molcel.2005.04.011. View

11.

Zang H, Goodenough A, Choi J, Irimia A, Loukachevitch L, Kozekov I . DNA adduct bypass polymerization by Sulfolobus solfataricus DNA polymerase Dpo4: analysis and crystal structures of multiple base pair substitution and frameshift products with the adduct 1,N2-ethenoguanine. J Biol Chem. 2005; 280(33):29750-64. DOI: 10.1074/jbc.M504756200. View

12.

Abbondanzieri E, Greenleaf W, Shaevitz J, Landick R, Block S . Direct observation of base-pair stepping by RNA polymerase. Nature. 2005; 438(7067):460-5. PMC: 1356566. DOI: 10.1038/nature04268. View

13.

Garcia-Diaz M, Bebenek K, Krahn J, Pedersen L, Kunkel T . Structural analysis of strand misalignment during DNA synthesis by a human DNA polymerase. Cell. 2006; 124(2):331-42. DOI: 10.1016/j.cell.2005.10.039. View

14.

Hariharan C, Bloom L, Helquist S, Kool E, Reha-Krantz L . Dynamics of nucleotide incorporation: snapshots revealed by 2-aminopurine fluorescence studies. Biochemistry. 2006; 45(9):2836-44. PMC: 2547141. DOI: 10.1021/bi051644s. View

15.

Kroeger K, Kim J, Goodman M, Greenberg M . Replication of an oxidized abasic site in Escherichia coli by a dNTP-stabilized misalignment mechanism that reads upstream and downstream nucleotides. Biochemistry. 2006; 45(15):5048-56. PMC: 1447609. DOI: 10.1021/bi052276v. View

16.

Mirkin E, Castro Roa D, Nudler E, Mirkin S . Transcription regulatory elements are punctuation marks for DNA replication. Proc Natl Acad Sci U S A. 2006; 103(19):7276-81. PMC: 1464333. DOI: 10.1073/pnas.0601127103. View

17.

Kunkel T, Bebenek K . DNA replication fidelity. Annu Rev Biochem. 2000; 69:497-529. DOI: 10.1146/annurev.biochem.69.1.497. View

18.

Huang J, Brieba L, Sousa R . Misincorporation by wild-type and mutant T7 RNA polymerases: identification of interactions that reduce misincorporation rates by stabilizing the catalytically incompetent open conformation. Biochemistry. 2000; 39(38):11571-80. DOI: 10.1021/bi000579d. View

19.

Suzuki M, Yoshida S, Adman E, Blank A, Loeb L . Thermus aquaticus DNA polymerase I mutants with altered fidelity. Interacting mutations in the O-helix. J Biol Chem. 2000; 275(42):32728-35. DOI: 10.1074/jbc.M000097200. View

20.

Mentesana P, Sousa R, McAllister W . Characterization of halted T7 RNA polymerase elongation complexes reveals multiple factors that contribute to stability. J Mol Biol. 2001; 302(5):1049-62. DOI: 10.1006/jmbi.2000.4114. View