Spatial relationship between polymerase and exonuclease active sites of phage T4 DNA polymerase enzyme

J Biol Chem. 1994 Aug 19;269(33):21123-6.

Abstract

The spatial relationship between the polymerase and exonuclease active sites of bacteriophage T4 DNA polymerase enzyme has been examined using a bulky biotin-streptavidin block at a specified position in an oligonucleotide (Fig. 1). The idea was to monitor the closest distance of approach of the T4 enzyme before being blocked by the bulky biotin-streptavidin complex while performing either of its activities. The results indicated a distance of 4-5 nucleotides between the biotin-streptavidin probe and the exonuclease site and a distance requirement of at least 7 nucleotides between the bulky probe and the 3'-primer terminus for efficient polymerization by the T4 enzyme. The difference in the two distances suggested a separation of 2-3 nucleotides between the two active sites of the T4 enzyme.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Bacterial Proteins
  • Base Sequence
  • Binding Sites
  • Biotin
  • DNA Primers
  • DNA-Directed DNA Polymerase / chemistry
  • DNA-Directed DNA Polymerase / metabolism*
  • Exodeoxyribonucleases / metabolism*
  • Molecular Sequence Data
  • Streptavidin
  • Viral Proteins / chemistry
  • Viral Proteins / metabolism*

Substances

  • Bacterial Proteins
  • DNA Primers
  • Viral Proteins
  • gene 43 protein, Enterobacteria phage T4
  • Biotin
  • Streptavidin
  • DNA-Directed DNA Polymerase
  • Exodeoxyribonucleases