title: Cleavage of the HIV-1 p66 reverse transcriptase/RNase H by the p9 protease in vitro generates active p15 RNase H.
abstract: The reverse transcriptase/RNase H of HIV-1 is composed of a p66/p51 heterodimer when analyzed from virus particles. A recombinant reverse transcriptase (RT)/RNase H which after purification consisted mainly of p66 was analyzed as substrate of the purified recombinant HIV-1 protease p9 in vitro. The p66 protein if treated with the protease is processed to a stable p66/p51 heterodimer. A p15 protein is a prominent cleavage product which was identified as the carboxyterminal portion of p66 by means of a monoclonal antibody. It exhibits RNase H activity when tested by activated gel analysis. Presence of SDS during the incubation allowed complete degradation of p66 depending on the conditions, which indicates that conformation of a substrate is relevant for cleavage by the HIV-1 protease. A synthetic heptapeptide AET-FYVD derived from the region between RT and RNase H is cleaved efficiently in vitro by the HIV-1 protease at the F'Y junction, and may mimick a natural cleavage site. P66/p51 heterodimers exhibit higher RT and RNase H activities than p66 when renatured from polyacrylamide gels.
authors.affiliation: Max-Planck-Institut für Molekulare Genetik, Berlin, Federal Republic of Germany.
authors.forename: T
authors.initials: T
authors.lastname: Schulze
authors.forename: M
authors.initials: M
authors.lastname: Nawrath
authors.forename: K
authors.initials: K
authors.lastname: Moelling
mesh_terms: D002851:Chromatography, High Pressure Liquid; D004722:Endoribonucleases / Q000378:metabolism*; D015684:Gene Products, pol / Q000378:metabolism*; D016333:HIV Protease / Q000378:metabolism; D015497:HIV-1 / Q000201:enzymology*; D009842:Oligopeptides / Q000138:chemical synthesis / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism*; D016914:Ribonuclease H; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology
chemical_list: D015684:Gene Products, pol; D009842:Oligopeptides; D012967:Sodium Dodecyl Sulfate; D012194:RNA-Directed DNA Polymerase; D004722:Endoribonucleases; D016914:Ribonuclease H; D016333:HIV Protease