Intermolecular binding sites of human immunodeficiency virus type 1 Rev protein determined by protein footprinting.

The Journal of Biological Chemistry
T H JensenJ Kjems

Abstract

Human immunodeficiency virus encodes the regulatory protein Rev, which is required for expression of viral structural proteins. It binds to an RNA element (RRE) in the viral transcript and up-regulates the cytoplasmic appearance of unspliced and singly spliced viral mRNA. We have studied the structure of Rev alone and complexed with the RRE and two monoclonal antibodies, using a protein footprinting approach. The method involves radioactive labeling at the C-terminal end of Rev fusion protein followed by limited proteolysis under native conditions, using 10 different proteinases. Rev protein was mainly cleaved within the basic domain and in the C-terminal part. The periodicity of the proteolytic cleavages within the basic domain strongly suggests that it forms an alpha-helical structure with one side facing the solvent. In the presence of RRE, these cleavages became significantly reduced. In addition, strong protection was observed at position 66 outside the basic domain. As a control for the specificity of the footprinting reaction, we confirmed the position of the epitopes for two monoclonal antibodies. This protein footprinting methodology is generally applicable to other proteins for which terminal modifications are accepta...Continue Reading

References

Sep 1, 1991·Proceedings of the National Academy of Sciences of the United States of America·M L ZappM R Green
Jan 15, 1992·Proceedings of the National Academy of Sciences of the United States of America·L S TileyB R Cullen
Feb 1, 1991·Proceedings of the National Academy of Sciences of the United States of America·J KjemsP A Sharp
Aug 10, 1990·Science·D D Chang, P A Sharp
Jun 1, 1990·Proceedings of the National Academy of Sciences of the United States of America·S DaeflerF Wong-Staal
Oct 1, 1990·Proceedings of the National Academy of Sciences of the United States of America·X B LuM L Hammarskjöld
Feb 1, 1990·Proceedings of the National Academy of Sciences of the United States of America·A W CochraneC A Rosen
Oct 1, 1985·Proceedings of the National Academy of Sciences of the United States of America·P MatsudairaE Atherton
Dec 7, 1989·Nature·M L Zapp, M R Green
Mar 1, 1989·Proceedings of the National Academy of Sciences of the United States of America·B K FelberG N Pavlakis
Jan 1, 1987·Annual Review of Biochemistry·A M EdelmanE G Krebs
Nov 25, 1993·Nucleic Acids Research·L GiverA D Ellington
Mar 15, 1994·Biochemistry·M AuerH Gstach
Jun 4, 1993·Cell·R TanA D Frankel

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Citations

Feb 13, 2001·European Journal of Biochemistry·A M Dirac, J Kjems
Jun 2, 1997·The EMBO Journal·J Lykke-AndersenJ Kjems
Aug 18, 2004·Journal of Molecular Biology·Birgitte H Kallipolitis, Poul Valentin-Hansen
Dec 5, 1998·The Journal of Biological Chemistry·P AskjaerJ Kjems
Oct 8, 1998·Journal of Molecular Biology·A JensenJ Kjems

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