An expanded model of replicating human immunodeficiency virus reverse transcriptase

Biochemistry. 1995 Apr 25;34(16):5343-56. doi: 10.1021/bi00016a005.


Replication complexes containing wild-type and RNase H-deficient p66/p51 human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) were analyzed by DNase I and S1 footprinting. While crystallography and chemical footprinting data demonstrate that 15-18 bases of primer and template occupy the DNA polymerase and RNase H active centers, enzymatic footprinting suggests that a larger portion of substrate is encompassed by the replicating enzyme. Independent of the position of DNA synthesis arrest, template nucleotides +7 to -23 and primer nucleotides -1 to -25 are nuclease resistant. On both DNA strands, position -20 remains accessible to DNase I cleavage, suggestive of an alteration in nucleic acid structure between exiting the RNase H catalytic center and leaving the C-terminal p66 domain. A model of HIV-1 RT containing an extended single-stranded template and duplex region was constructed on the basis of the structure of an RT/DNA complex. Mapping of footprint data onto this model shows consistency between biochemical and structural data, implicating a contribution from domains proximal to the catalytic centers.

Publication types

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

MeSH terms

  • Base Sequence
  • Binding Sites
  • DNA Primers
  • DNA Replication*
  • DNA, Viral / biosynthesis
  • DNA, Viral / chemistry*
  • DNA-Directed DNA Polymerase / metabolism
  • Deoxyribonuclease I
  • HIV Reverse Transcriptase
  • HIV-1 / enzymology*
  • Models, Molecular
  • Molecular Sequence Data
  • Nucleic Acid Conformation
  • Protein Conformation*
  • Protein Structure, Secondary
  • RNA-Directed DNA Polymerase / biosynthesis
  • RNA-Directed DNA Polymerase / chemistry*
  • RNA-Directed DNA Polymerase / metabolism*
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Ribonuclease H / metabolism
  • Substrate Specificity
  • Templates, Genetic


  • DNA Primers
  • DNA, Viral
  • Recombinant Proteins
  • HIV Reverse Transcriptase
  • RNA-Directed DNA Polymerase
  • DNA-Directed DNA Polymerase
  • Deoxyribonuclease I
  • Ribonuclease H