Untitled

Abstract

We have analyzed the folding pathway of the tetramerization domain of the tumor suppressor protein p53. Structures of transition states were determined from phi-values for 25 mutations, including leucine to norvaline, and the analysis encompassed nearly every residue in the domain. Denatured monomers fold and dimerize, through a transition state with little native structure, to form a transient, highly structured dimeric intermediate. The intermediate dimerizes, through a native-like transition state with the primary dimers fully folded but with interdimer interactions only partially formed, to form the native tetramer as a 'dimer of dimers'.

Single-Molecule characterization of oligomerization kinetics and equilibria of the tumor suppressor p53

“…The rate constant for the association of p53tet at 25°C was previously measured from protein folding studies under somewhat different conditions to be 3.1 × 10 5  s −1 M −1 , and the dissociation rate constant for p53tet tetramer 3.7 × 10 −3 s −1 (13), which are in reasonable agreement with values in Table 3 (4.2 × 10 5 s −1 and 1.8 × 10 −3  s −1 , respectively). The rate constants for the association of p53tet are 2- to 3-fold higher than for full-length p53.…”

“…Moreover, post-translational modifications such as phosphorylation, acetylation and ubiquitination sometimes require p53 to be tetramerized (10–12). Experimentally, formation of tetrameric p53 occurs by the following equilibrium 4M↔ 2D ↔ T (where M = monomer, D = dimer and T = tetramer) (13). Furthermore, p53 dimerizes on co-translation in polysomes but tetramerization occurs post-translationally in solution (4).…”

Cotranslational protein assembly imposes evolutionary constraints on homomeric proteins

“…First, the Tet oligomerization domain has a low (~nM) dissociation constant24, in line with in vivo observations that Tet exists as oligomers14. Second, Tet folds and assembles faster than its translation rate12, which means it is expected to fold soon after it exits the ribosome tunnel, as observed previously13. Therefore, if positioned at the N-terminus, Tet is likely to oligomerize during translation of the reporter domain.…”

“…(Sequences provided in Supplementary Note). Thus, their translation takes orders of magnitude longer than Tet folding-12,16,17, considering that bacterial translation rates are on the order of 10-20 amino acids per second18.…”

Pathological Unfoldomics of Uncontrolled Chaos: Intrinsically Disordered Proteins and Human Diseases

“…This view is also supported by studies of p53 tetramerization domain folding, which showed that the initial assembly is between highly unstructured monomers that form a transient, highly structured dimeric intermediate. 171 …”