DNA transcription is initiated by a small regulatory region of transactivators known as the transactivation domain. In contrast to the rapid progress made on the functional aspect of this promiscuous domain, its structural feature is still poorly characterized. Here, our multidimensional NMR study reveals that an unbound fulllength p53 transactivation domain, although similar to the recently discovered group of loosely folded proteins in that it does not have tertiary structure, is nevertheless populated by an amphipathic helix and two nascent turns.

“What’s in a name? That which we call a rose By any other name would smell as sweet.” From “Romeo and Juliet”, William Shakespeare (1594) This article opens a series of publications on disambiguation of the basic terms used in the field of intrinsically disordered proteins. We start from the beginning, namely from the explanation of what the expression “intrinsically disordered protein” actually means and why this particular term has been chosen as the common denominator for this class of proteins characterized by broad structural, dynamic and functional characteristics.

Mdm2 is a cellular antagonist of p53 that keeps a balanced cellular level of p53. The two proteins are linked by a negative regulatory feedback loop and physically bind to each other via a putative helix formed by residues 18 -26 of p53 transactivation domain (TAD) and its binding pocket located within the N-terminal 100-residue domain of mdm2 (Kussie, P. H., Gorina, S., Marechal, V., Elenbaas, B., Moreau, J., Levine, A. J., and Pavletich, N. P. (1996) Science 274, 948 -953). In a previous report we demonstrated that p53 TAD in the mdm2-freee state is mostly unstructured but contains two nascent turns in addition to a "preformed" helix that is the same as the putative helix mediating p53-mdm2 binding. Here, using heteronuclear multidimensional NMR methods, we show that the two nascent turn motifs in p53 TAD, turn I (residues 40 -45) and turn II (residues 49 -54), are also capable of binding to mdm2. In particular, the turn II motif has a higher mdm2 binding affinity (ϳ20 M) than the turn I and targets the same site in mdm2 as the helix. Upon mdm2 binding this motif becomes a well defined full helix turn whose hydrophobic face formed by the side chains of Ile-50, Trp-53, and Phe-54 inserts deeply into the helix binding pocket. Our results suggest that p53-mdm2 binding is subtler than previously thought and involves global contacts such as multiple "non-contiguous" minimally structured motifs instead of being localized to one small helix mini-domain in p53 TAD.p53 is known to be implicated in more than 50% of all human cancers and probably represents one of the proteins that is most critically associated with cancer (1, 2). Understanding how this "hub" in the cancer protein network interacts with other members of the network is not only important for gaining insights into fundamental principles underlying tumorigenesis but also for efficient development of anticancer agents (3-6). Ironically, establishing a structure-function relationship for p53 has been possible only for ϳ30% of its amino acid residues; namely, for those forming globular domains, a DNA binding domain (7) and an oligomerization domain (8, 9). This fact can be attributed to a rather interesting finding that a large fraction (ϳ70%) of amino acid residues in p53 does not participate in forming a well defined tertiary structure, a common feature shared by many intrinsically unstructured proteins (IUPs) 2 (10 -16).IUPs are an interesting class of proteins that maintain their function despite the lack of a well defined globular structure. Structurally, IUPs are in a similar state as folding intermediates but are distinct from the latter in that they are not in an artificially denatured state. Although some IUPs totally lack any structural elements, others have minimal secondary structural elements (12,16,(17)(18)(19). One subgroup of IUPs consists of unstructured or flexible domains consisting of more than ϳ50 amino acid residues within large mother proteins (12,20,21). Because of their flexible nature, structural features of IUPs can be characterized ...

Intrinsically unfolded proteins (IUPs) do not obey the golden rule of structural biology, 3D structure = function, as they manifest their inherent functions without resorting to three-dimensional structures. Absence of a compact globular topology in these proteins strongly implies that their ligand recognition processes should involve factors other than spatially well-defined binding pockets. Heteronuclear multidimensional (HetMulD) NMR spectroscopy assisted with a stable isotope labeling technology is a powerful tool for quantitatively investigating detailed structural features in IUPs. In particular, it allows us to delineate the presence and locations of pre-structured motifs (PreSMos) on a per-residue basis. PreSMos are the transient local structural elements that presage target-bound conformations and act as specificity determinants for IUP recognition by target proteins. Here, we present a brief chronicle of HetMulD NMR studies on IUPs carried out over the past two decades along with a discussion on the functional significance of PreSMos in IUPs.