Trifluoroethanol Stabilizes a Helix‐Turn‐Helix Motif in Equine Infectious‐Anemia‐Virus Trans ‐Activator Protein

The solution structure of the 75‐amino‐acid trans ‐activator (Tat) protein of the equine infectious‐anemia virus in trifluoroethanol‐containing solution was determined by two‐dimensional and three‐dimensional nuclear magnetic resonance spectroscopy, resulting in a total of 838 nuclear‐Over‐hauser‐enhancement distance restraints, and restrained molecular‐dynamics simulations. In contrast to the recently determined structure of this protein in trifluoroethanol‐free pH 6.3 solution, the hydrophobic core and the adjacent basic RNA‐binding region of the protein showed well‐defined α‐helical secondary structure in trifluoroethanol‐containing solution. The helical regions comprise those parts of the molecule whose helix‐forming tendencies were noted earlier in trifluoroethanol‐free solution. Two helices (Gln38–Arg43 and Asp48–Ala64) are connected by a tight type‐II turn centered at the strictly conserved Gly46 leading to a helix‐turn‐helix motif in the core and basic region of the protein. A third helix (Thr9–Asn13) is located in the less well defined N‐terminal part of the protein. These observations may support the notion that the protein adopts a helical structure in the RNA‐binding region on complex formation. Although the secondary‐structure elements become better defined in trifluoroethanol‐containing solution, the opposite is true for the hydrophobically stabilized tertiary structure. This adds a caveat to studies of protein structures in trifluoroethanol‐containing solution in general.