Expression of the C-terminus of HIV-1 reverse transcriptase p66 and p51 subunits as a single polypeptide with RNase H activity.

Abstract

The C-terminus of the HIV-1 reverse transcriptase heterodimer was reconstructed into a single polypeptide. The construct encodes the p51 thumb (T) and connection (C) subdomains joined through a linker region to the p66 connection (C) and RNase H (R) domain. The TCCR protein was purified from insoluble fractions of Escherichia coli lysates. The TCCR construct maintains Mn(2+)-dependent RNase H activity and specifically cleaves the substrate mimicking the tRNA removal required for second-strand transfer reactions.