Temperature inducible beta-sheet structure in the transactivation domains of retroviral regulatory proteins of the Rev family

The interaction of the human immunodeficiency virus type 1 (HIV-I) regulato ry protein Rev with cellular cofactors is crucial for the viral life cycle. The HIV-1 Rev transactivation domain is functionally interchangeable with analog regions of Rev proteins of other retroviruses suggesting common fold ing patterns. In order to obtain experimental evidence for similar structur al features mediating protein-protein contacts we investigated activation d omain peptides from HIV-1, HIV-2, VISNA virus, feline immunodeficiency viru s (FIV) and equine infectious anemia virus (EIAV) by CD spectroscopy, secon dary structure prediction and sequence analysis. Although different in pola rity and hydrophobicity, all peptides showed a similar behavior with respec t to solution conformation, concentration dependence and variations in ioni c strength and pH. Temperature studies revealed an unusual induction of bet a-structure with rising temperatures in all activation domain peptides. The high stability of beta-structure in this region was demonstrated in three different peptides of the activation domain of HIV-I Rev in solutions conta ining 40% hexafluoropropanol, a reagent usually known to induce cc-helix in to amino acid sequences. Sequence alignments revealed similarities between the polar effector domains from FIV and EIAV and the leucine rich (hydropho bic) effector domains found in HIV-1, HIV-2 and VISNA. Studies on activatio n domain peptides of two dominant negative HIV-1 Rev mutants, M10 and M32, pointed towards different reasons for the biological behavior. Whereas the peptide containing the M10 mutation ((LE79)-E-78 --> (DL79)-L-78) showed wi ld-type structure, the M32 mutant peptide ((LLL83)-L-78-L-81 --> A(78)A(81) A(83)) revealed a different protein fold to be the reason for the disturbed binding to cellular cofactors. From our data, we conclude, that the activa tion domain of Rev proteins from different viral origins adopt a similar fo ld and that a beta-structural element is involved in binding to a cellular cofactor. (C) 1999 Elsevier Science B.V. All rights reserved.