Cleavage of the HIV-1 p66 reverse transcriptase/RNase H by the p9 protease in vitro generates active p15 RNase H

• Published in: Archives of virology 1991, Vol.118 (3-4), p.179-188
• Main Authors: Schulze, T, Nawrath, M, Moelling, K
• Format: Article
• Language: English
• Subjects: AIDS/HIV; Chromatography, High Pressure Liquid; Endoribonucleases - metabolism; Gene Products, pol - metabolism; HIV Protease - metabolism; HIV-1 - enzymology; human immunodeficiency virus 1; Oligopeptides - chemical synthesis; Oligopeptides - metabolism; Ribonuclease H; RNA-Directed DNA Polymerase - metabolism; Sodium Dodecyl Sulfate - pharmacology
• ISSN: 0304-8608; 1432-8798
• DOI: 10.1007/BF01314028

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.