Reverse transcriptase from Escherichia coli exists as a complex with msDNA and is able to synthesize double-stranded DNA

J Biol Chem. 1990 May 25;265(15):8490-6.

Abstract

Reverse transcriptase required for the synthesis of msDNA.Ec67 in an Escherichia coli strain was purified as a large molecular weight complex with msDNA. The complex sedimented in a glycerol gradient at an s value greater than 19. The predominant protein species co-purifying with reverse transcriptase activity in the complex had a molecular weight estimated at 65,000 which is close to the expected size of 67,227 for the Ec67-reverse transcriptase. In addition, the large complex also contained msDNA.Ec67. The purified complex was able to synthesize cDNA using 5 S rRNA as a template (annealed to a synthetic DNA primer), and a double-stranded DNA using a synthetic DNA template (annealed to a synthetic DNA primer). When msDNA.Ec67 was used as a natural template:primer, the purified complex produced two major products: a 103-base single-stranded DNA by extending the 3' end of msDNA using msdRNA as a template, and a 60-base double-stranded DNA product resulting from the converse reaction in which the 3' end of msdRNA is extended using msDNA as a template. The results suggest that bacterial reverse transcriptase is capable of producing single-stranded cDNA and possibly double-stranded DNA as well. Possible implications of these findings on the biology of the msDNA-retron system are discussed.

Publication types

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

MeSH terms

  • Base Sequence
  • DNA, Bacterial / biosynthesis*
  • DNA, Single-Stranded / biosynthesis
  • DNA, Single-Stranded / isolation & purification
  • DNA, Single-Stranded / metabolism*
  • Escherichia coli / enzymology*
  • Macromolecular Substances
  • Molecular Sequence Data
  • Molecular Weight
  • Nucleic Acid Conformation
  • Oligonucleotide Probes
  • Plasmids
  • RNA, Bacterial / biosynthesis
  • RNA, Bacterial / isolation & purification
  • RNA, Bacterial / metabolism*
  • RNA, Ribosomal, 5S / genetics
  • RNA-Directed DNA Polymerase / isolation & purification
  • RNA-Directed DNA Polymerase / metabolism*
  • Restriction Mapping
  • Templates, Genetic

Substances

  • DNA, Bacterial
  • DNA, Single-Stranded
  • Macromolecular Substances
  • Oligonucleotide Probes
  • RNA, Bacterial
  • RNA, Ribosomal, 5S
  • msDNA
  • RNA-Directed DNA Polymerase