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Misincorporation and mispaired primer extension by human immunodeficiency virus reverse transcriptase.

Publication ,  Journal Article
Zinnen, S; Hsieh, JC; Modrich, P
Published in: J Biol Chem
September 30, 1994

Pre-steady-state methods were used to study the fidelity of human immunodeficiency virus reverse transcriptase. Fidelity of DNA-directed DNA synthesis can be attributed to a 1-2 order of magnitude reduction in affinity for noncomplementary dNTPs, and a 1-4 order of magnitude reduction in the rate of the conformational change that limits the rate of nucleotide addition. Affinities of reverse transcriptase for paired or mispaired primer termini are similar. Discrimination against a mispaired primer is due to reduction in affinity for the next dNTP and reduction in rate of extension. Extension of mispaired termini proceeds 20-700-fold faster than the rate of dissociation of reverse transcriptase from the primer-template and is 2-3 orders of magnitude more frequent than nucleotide misincorporation. The rate-limiting step for extension of a mispaired terminus occurs at the conformational change or chemical step, depending on the nature of the mispair. Presence of a mismatch at the 3' penultimate position reduces pyrophosphorolysis of the primer by a factor of 10(3), indicating that mispairs 5' to the site of chemistry can also affect catalysis.

Duke Scholars

Published In

J Biol Chem

ISSN

0021-9258

Publication Date

September 30, 1994

Volume

269

Issue

39

Start / End Page

24195 / 24202

Location

United States

Related Subject Headings

  • Templates, Genetic
  • RNA-Directed DNA Polymerase
  • Protein Conformation
  • Protein Binding
  • Nucleotides
  • Nucleic Acid Heteroduplexes
  • Molecular Sequence Data
  • Kinetics
  • HIV
  • DNA Replication
 

Citation

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Zinnen, S., Hsieh, J. C., & Modrich, P. (1994). Misincorporation and mispaired primer extension by human immunodeficiency virus reverse transcriptase. J Biol Chem, 269(39), 24195–24202.
Zinnen, S., J. C. Hsieh, and P. Modrich. “Misincorporation and mispaired primer extension by human immunodeficiency virus reverse transcriptase.J Biol Chem 269, no. 39 (September 30, 1994): 24195–202.
Zinnen S, Hsieh JC, Modrich P. Misincorporation and mispaired primer extension by human immunodeficiency virus reverse transcriptase. J Biol Chem. 1994 Sep 30;269(39):24195–202.
Zinnen, S., et al. “Misincorporation and mispaired primer extension by human immunodeficiency virus reverse transcriptase.J Biol Chem, vol. 269, no. 39, Sept. 1994, pp. 24195–202.
Zinnen S, Hsieh JC, Modrich P. Misincorporation and mispaired primer extension by human immunodeficiency virus reverse transcriptase. J Biol Chem. 1994 Sep 30;269(39):24195–24202.

Published In

J Biol Chem

ISSN

0021-9258

Publication Date

September 30, 1994

Volume

269

Issue

39

Start / End Page

24195 / 24202

Location

United States

Related Subject Headings

  • Templates, Genetic
  • RNA-Directed DNA Polymerase
  • Protein Conformation
  • Protein Binding
  • Nucleotides
  • Nucleic Acid Heteroduplexes
  • Molecular Sequence Data
  • Kinetics
  • HIV
  • DNA Replication