An Extended α-Helix and Specific Amino Acid Residues Opposite the DNA-binding Surface of the cAMP Response Element Binding Protein Basic Domain Are Important for Human T Cell Lymphotropic Retrovirus Type I Tax Binding

Abstract

The human T cell lymphotropic retrovirus type I (HTLV-I) trans-activator, Tax, interacts specifically with the basic-domain/leucine-zipper (bZip) protein, cAMP response element binding protein (CREB), bound to the viral Tax-responsive element consisting of three imperfect 21-base pair repeats, each with a cAMP response element core flanked by G/C-rich sequences. Here, the minimal CREB-bZip necessary for Tax binding is shown to be composed of amino acid residues 280-341. The Tax-CREB interaction involves an uninterrupted and extended alpha-helix in CREB that spans most of its basic domain to include amino acid residues localized to the NH2 terminus of the DNA binding region. Mutational analyses indicate that three residues, Arg284, Met291, and Glu299 unique to this region of the CREB/activating transcription factor-1 subfamily of bZip proteins, constitute the contact surface for Tax. Amino acid substitutions in these positions had little impact on CREB-bZip binding to DNA but abrogated its binding to Tax. Each of the contact residues for Tax are spaced approximately two helical turns apart on the side of the bZip helix directly opposite to that of the invariant DNA-binding residues. Molecular modeling reveals the Tax-contact residues to be near the minor groove of the G/C-rich DNA in the 21-base pair repeat. They most likely position Tax for minor groove contact with the G/C-rich sequences.