Isolation and characterization of a split B-type DNA polymerase from the archaeon Methanobacterium thermoautotrophicum deltaH

J Biol Chem. 1999 Oct 1;274(40):28751-61. doi: 10.1074/jbc.274.40.28751.


We describe here the isolation and characterization of a B-type DNA polymerase (PolB) from the archaeon Methanobacterium thermoautotrophicum DeltaH. Uniquely, the catalytic domains of M. thermoautotrophicum PolB are encoded from two different genes, a feature that has not been observed as yet in other polymerases. The two genes were cloned, and the proteins were overexpressed in Escherichia coli and purified individually and as a complex. We demonstrate that both polypeptides are needed to form the active polymerase. Similar to other polymerases constituting the B-type family, PolB possesses both polymerase and 3'-5' exonuclease activities. We found that a homolog of replication protein A from M. thermoautotrophicum inhibits the PolB activity. The inhibition of DNA synthesis by replication protein A from M. thermoautotrophicum can be relieved by the addition of M. thermoautotrophicum homologs of replication factor C and proliferating cell nuclear antigen. The possible roles of PolB in M. thermoautotrophicum replication are discussed.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • Chromatography, Affinity
  • Cloning, Molecular
  • DNA Polymerase beta / genetics
  • DNA Polymerase beta / isolation & purification*
  • DNA Polymerase beta / metabolism
  • DNA Replication
  • DNA-Binding Proteins / metabolism
  • Electrophoresis, Polyacrylamide Gel
  • Methanobacterium / enzymology*
  • Methanobacterium / genetics
  • Molecular Sequence Data
  • Oligonucleotides
  • Recombinant Proteins / genetics
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism


  • DNA-Binding Proteins
  • Oligonucleotides
  • Recombinant Proteins
  • DNA Polymerase beta

Associated data

  • GENBANK/AE000901