Interactions of Escherichia coli thioredoxin, the processivity factor, with bacteriophage T7 DNA polymerase and helicase

J Biol Chem. 2008 Nov 14;283(46):32077-84. doi: 10.1074/jbc.M805062200. Epub 2008 Aug 30.


Escherichia coli thioredoxin binds to a unique flexible loop of 71 amino acid residues, designated the thioredoxin binding domain (TBD), located in the thumb subdomain of bacteriophage T7 gene 5 DNA polymerase. The initial designation of thioredoxin as a processivity factor was premature. Rather it remodels the TBD for interaction with DNA and the other replication proteins. The binding of thioredoxin exposes a number of basic residues on the TBD that lie over the duplex region of the primer-template and increases the processivity of nucleotide polymerization. Two small solvent-exposed loops (loops A and B) located within TBD electrostatically interact with the acidic C-terminal tail of T7 gene 4 helicase-primase, an interaction that is enhanced by the binding of thioredoxin. Several basic residues on the surface of thioredoxin in the polymerase-thioredoxin complex lie in close proximity to the TBD. One of these residues, lysine 36, is located proximal to loop A. The substitution of glutamate for lysine has a dramatic effect on the binding of gene 4 helicase to a DNA polymerase-thioredoxin complex lacking charges on loop B; binding is decreased 15-fold relative to that observed with wild-type thioredoxin. This defective interaction impairs the ability of T7 DNA polymerase-thioredoxin together with T7 helicase to mediate strand displacement synthesis. This is the first demonstration that thioredoxin interacts with replication proteins other than T7 DNA polymerase.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Bacteriophage T7 / enzymology*
  • Bacteriophage T7 / genetics
  • Crystallography, X-Ray
  • DNA / biosynthesis
  • DNA Helicases / chemistry*
  • DNA Helicases / genetics
  • DNA Helicases / metabolism*
  • DNA-Directed DNA Polymerase / chemistry*
  • DNA-Directed DNA Polymerase / genetics
  • DNA-Directed DNA Polymerase / metabolism*
  • Escherichia coli / chemistry
  • Escherichia coli / genetics
  • Escherichia coli / metabolism*
  • Models, Molecular
  • Mutation / genetics
  • Protein Binding
  • Protein Structure, Quaternary
  • Substrate Specificity
  • Thioredoxins / chemistry
  • Thioredoxins / genetics
  • Thioredoxins / metabolism*


  • Thioredoxins
  • DNA
  • bacteriophage T7 induced DNA polymerase
  • DNA-Directed DNA Polymerase
  • DNA Helicases