title: Formation of heterodimers of human-immunodeficiency-virus-type-1 reverse transcriptase by recombination of separately purified subunits.
abstract: Human-immunodeficiency-virus-type-1 reverse transcriptase exists in virions as a heterodimer of a M(r) 66,000 subunit and its C-terminally truncated form of M(r) 51,000, but, when expressed as a recombinant M(r) 66,000 protein, a mixture of heterodimers and homodimers results which co-purify by most conventional techniques. We describe a method of hydrophobic chromatography which gives baseline separation of these two forms of the protein. This method has been applied to purify heterodimers formed by recombination of separately expressed and purified M(r) 66,000 and 51,000 subunits, resulting in significantly more homogeneous heterodimer preparations. The recombined heterodimer showed similar kinetic properties and RNase H activity to the standard heterodimer and a specific activity significantly higher than the original homodimer of the M(r) 66,000 protein. Heterodimers having greater asymmetry have also been prepared by recombining Mr 66,000 subunits containing single-point or deletion mutations, with wild-type M(r) 51,000 subunits, and the resulting heterodimers analysed.
authors.affiliation: Department of Molecular Sciences, Wellcome Research Laboratories, Beckenham, Kent, England.
authors.forename: D K
authors.initials: DK
authors.lastname: Stammers
authors.forename: C K
authors.initials: CK
authors.lastname: Ross
authors.forename: H
authors.initials: H
authors.lastname: Idriss
authors.forename: D M
authors.initials: DM
authors.lastname: Lowe
mesh_terms: D002850:Chromatography, Gel; D002853:Chromatography, Liquid; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D015497:HIV-1 / Q000201:enzymology*; D007700:Kinetics; D009154:Mutation; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016914:Ribonuclease H / Q000378:metabolism
chemical_list: D012194:RNA-Directed DNA Polymerase; D016914:Ribonuclease H