HIV-1 reverse transcriptase is phosphorylated in vitro and in a cellular system.
Phosphorylation modulates the activity of many proteins that interact with nucleic acids including DNA and RNA polymerases. The HIV-1 reverse transcriptase (RT) is essential during the replicative cycle of the HIV-1 virus. HIV-1 RT has several potential sites for phosphorylation that could regulate its activities. In this work, the phosphorylation of HIV-1 RT is examined in vitro and in vivo, to evaluate any role for this modification in regulating RT metabolism. Recombinant unphosphorylated HIV-1 RT heterodimer expressed in bacteria can be phosphorylated in vitro by several purified mammalian protein kinases. Seven kinases were tested, and five of these enzymes phosphorylated HIV-1 RT. Using an insect baculovirus expression system, the 66 kDa HIV-1 RT was also phosphorylated in vivo. However, HIV-1 RT immunoprecipitated from H9-lymphoma cells infected with HIV-1 showed negligible phosphorylation. Our results indicate that purified HIV-1 RT can be phosphorylated by several mammalian protein kinases in vitro and during expression in baculovirus infected insect cells.
Idriss
H
H
Sealy Center for Molecular Science, The University of Texas Medical Branch Galveston, TX 77555-0851, USA. hi@st-and.ac.uk
Kawa
S
S
Damuni
Z
Z
Thompson
E B
EB
Wilson
S H
SH
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D002460:Cell Line; D054303:HIV Reverse Transcriptase / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D010710:Phosphates / Q000378:metabolism; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D018411:Spodoptera; D013379:Substrate Specificity
D000911:Antibodies, Monoclonal; D010710:Phosphates; D010761:Phosphorus Radioisotopes; D011994:Recombinant Proteins; D011494:Protein Kinases; D054303:HIV Reverse Transcriptase