Expression of the C-terminus of HIV-1 reverse transcriptase p66 and p51 subunits as a single polypeptide with RNase H activity.
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.
Zúñiga
Roberto
R
Programa de Virología, Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, Independencia 1027, Santiago, Chile.
Sengupta
Sonali
S
Snyder
Christine
C
Leon
Oscar
O
Roth
Monica J
MJ
D015153:Blotting, Western; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D054303:HIV Reverse Transcriptase / Q000737:chemistry*; D008345:Manganese / Q000737:chemistry; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods; D017434:Protein Structure, Tertiary; D012343:RNA, Transfer / Q000737:chemistry; D016914:Ribonuclease H / Q000737:chemistry*; D012685:Sepharose / Q000737:chemistry; D012984:Software; D014508:Urea / Q000737:chemistry
D010455:Peptides; D008345:Manganese; D014508:Urea; D012685:Sepharose; D012343:RNA, Transfer; D054303:HIV Reverse Transcriptase; D016914:Ribonuclease H