TRANSCRIPTIONAL ACTIVATOR-COACTIVATOR RECOGNITION - NASCENT FOLDING OF A KINASE-INDUCIBLE TRANSACTIVATION DOMAIN PREDICTS ITS STRUCTURE ON COACTIVATOR BINDING

A model of transcriptional activator-coactivator recognition is provid ed by the mammalian CREB activation domain and the KM domain of coacti vator CBP. The CREB kinase-inducible activation domain (pKID, 60 resid ues) is disordered in solution and undergoes an a-helical folding tran sition on binding to CBP [Radhakrishan, I., Perez-Alvarado, G. C., Par ker, D., Dyson, H. J., Montminy, M. R., and Wright, P. E. (1997) Cell 91, 741-752]. Binding requires phosphorylation of a conserved serine ( RPpSYR) in pKID associated in vivo with the biological activation of C REB signaling pathways. The CBP-bound structure of CREB contains two a lpha-helices (designated alpha A and alpha B) flanking the phosphoseri ne; the bound structure is stabilized by specific interactions with CB P. Here, the nascent structure of an unbound pKID domain is characteri zed by multidimensional NMR spectroscopy. The solubility of the phosph opeptide (46 residues) was enhanced by truncation of N- and C-terminal residues not involved in pKID-CBP interactions. Although disordered u nder physiologic conditions, the pKID fragment and its unphosphorylate d parent peptide exhibit partial folding at low temperatures. One reco gnition helix (alpha A) is well-defined at 4 degrees C, whereas the ot her (alpha B) is disordered but inducible in 40% trifluoroethanol (TFE ). Such nascent structure is independent of serine phosphorylation and correlates with the relative extent of engagement of the two alpha-he lices in the pKID-KIX complex; whereas alpha A occupies a peripheral b inding site with few intermolecular contacts, the TFE-inducible alpha B motif is deeply engaged in a hydrophobic groove. Our results support the use of TFE as an empirical probe of hidden structural propensitie s and define a correspondence between induced fit and the nascent stru cture of peptide fragments.