Purification and characterization of the RNase H domain of HIV-1 reverse transcriptase expressed in recombinant Escherichia coli

FEBS Lett. 1990 Sep 17;270(1-2):76-80. doi: 10.1016/0014-5793(90)81238-j.


The ribonuclease H (RNase H) domain of human immuno-deficiency virus (HIV-1) reverse transcriptase has been produced with the aim of providing sufficient amounts of protein for biophysical studies. A plasmid vector is described which directs high level expression of the RNase H domain under the control of the lambda PL promoter. The domain corresponds to residues 427-560 of the 66 kDa reverse transcriptase. The protein was expressed in Escherichia coli and was purified using ion-exchange and size exclusion chromatography. The purified protein appears to be in a native-like homogeneous conformational state as determined by 1H-NMR spectroscopy and circular dichroism measurements. HIV-protease treatment of the RNase H domain resulted in cleavage between Phe-440 and Tyr-441.

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
  • Endopeptidases
  • Endoribonucleases / biosynthesis
  • Endoribonucleases / chemistry*
  • Escherichia coli / metabolism
  • HIV-1 / enzymology*
  • Magnetic Resonance Spectroscopy
  • Molecular Sequence Data
  • Molecular Weight
  • Peptide Fragments / chemistry
  • Protein Conformation
  • RNA-Directed DNA Polymerase / chemistry*
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / chemistry
  • Ribonuclease H
  • Structure-Activity Relationship
  • X-Ray Diffraction


  • Peptide Fragments
  • Recombinant Proteins
  • RNA-Directed DNA Polymerase
  • Endoribonucleases
  • Ribonuclease H
  • Endopeptidases