Differences in the transactivation domains of p53 family members: a computational study

Abstract

The N terminal transactivation domain of p53 is regulated by ligases and coactivator proteins. The functional conformation of this region appears to be an alpha helix which is necessary for its appropriate interactions with several proteins including MDM2 and p300. Folding simulation studies have been carried out to examine the propensity and stability of this region and are used to understand the differences between the family members with the ease of helix formation following the order p53 > p73 > p63. It is clear that hydrophobic clusters control the kinetics of helix formation, while electrostatic interactions control the thermodynamic stability of the helix. Differences in these interactions between the family members may partially account for the differential binding to, and regulation by, MDM2 (and MDMX). Phosphorylations of the peptides further modulate the stability of the helix and control associations with partner proteins.

Figure 1

Folding pattern of p53 (A) Evolution of secondary structures of the p53 peptides as a function of simulation time Colour code: purple, α-helix; red, π-helix; yellow, β-sheet; green, isolated bridge; cyan, turn; white, random coil. (B) Hydrogen bond statistics of the secondary structures averaged over 100 ns of simulations; the lifetime of hydrogen bonds in 5 ns windows is shown as: Space ( ) for 0-5%, dot (.) for 5-20%, dash (-) for 20-40%, o for 40-60%, x for 60-80%, star (*) for 80-95% and at (@) for 95 – 100%. (C) Cluster analysis of secondary structures in terms of RMSD as a function of simulation time; a representative structure (N-terminus in blue, C-terminus in red) from each cluster is shown with % of population; colour code of the plot: red is helix, yellow is β-Sheet and green is random structure. Conserved residues F19, W23 and L26 are shown as sticks. (D) Snapshot of the putative nucleation conformation of p53during the folding simulation; nucleation residue L22 and hydrogen bond between D21 side chain and backbone of W23 are shown. Hydrogen Bonds are shown as red dotted lines.

Figure 2

Folding pattern of p63 (A) Evolution of secondary structures of the p63 peptides as a function of simulation time Colour code: purple, α-helix; red, π-helix; yellow, β-sheet; green, isolated bridge; cyan, turn; white, random coil. (B) Hydrogen bond statistics of the secondary structures averaged over 100 ns of simulations; the lifetime of hydrogen bonds in 5 ns windows is shown as: Space ( ) for 0-5%, dot (.) for 5-20%, dash (-) for 20-40%, o for 40-60%, x for 60-80%, star (*) for 80-95% and at (@) for 95 – 100%. (C) Cluster analysis of secondary structures in terms of RMSD as a function of simulation time; a representative structure (N-terminus in blue, C-terminus in red) from each cluster is shown with % of population; colour code of the plot: red is helix, yellow is β-Sheet and green is random structure. Conserved residues F19, W23 and L26 are shown as sticks.

Figure 3

Folding pattern of p73 Evolution of secondary structures of the p73 peptides as a function of simulation time Colour code: purple, α-helix; red, π-helix; yellow, β-sheet; green, isolated bridge; cyan, turn; white, random coil. (B) Hydrogen bond statistics of the secondary structures averaged over 100 ns of simulations; the lifetime of hydrogen bonds in 5 ns windows is shown as: Space ( ) for 0-5%, dot (.) for 5-20%, dash (-) for 20-40%, o for 40-60%, x for 60-80%, star (*) for 80-95% and at (@) for 95 – 100%. (C) Cluster analysis of secondary structures in terms of RMSD as a function of simulation time; a representative structure (N-terminus in blue, C-terminus in red) from each cluster is shown with % of population; colour code of the plot: red is helix, yellow is β-Sheet and green is random structure. Conserved residues F19, W23 and L26 are shown as sticks.

Figure 4

Evolution of secondary structures of the peptide variants at position 22 along the simulation: (A) p53: L22I; (B) p63: I22L; (C) p73: L22I; Colour code: purple, α-helix; red, π-helix; yellow, β-sheet; green, isolated bridge; cyan, turn; white, random coil.

Figure 5

Evolution of secondary structures of the peptide variants at position 21 and 24 along the simulation: (A) p63: H21D, D24K; (B) p73: H21D, S24K; Colour code: purple, α-helix; red, π-helix; yellow, β-sheet; green, isolated bridge; cyan, turn; white, random coil.

Figure 6

Evolution of secondary structures of the peptide variants at position 21, 22 and 24 along the simulation; Colour code: purple, α-helix; red, π-helix; yellow, β-sheet; green, isolated bridge; cyan, turn; white, random coil.

Figure 7

Evolution of secondary structures of the peptide variants at position 25 along the simulation: (A) p53: L25F (B) p63: F25L (C) p73: S25F; Colour code: purple, α-helix; red, π-helix; yellow, β-sheet; green, isolated bridge; cyan, turn; white, random coil.

Figure 8

Evolution of secondary structures of the phosphorylated peptide variants of p53 at (A) T18 (B) S20 and (C) T18 and S20; Colour code: purple, α-helix; red, π-helix; yellow, β-sheet; green, isolated bridge; cyan, turn; white, random coil.

Figure 9

Evolution of secondary structures of the phosphorylated peptide variants of p73 at (A) T17 (B) T18 (C) S24 and (D) S25; Colour code: purple, α-helix; red, π-helix; yellow, β-sheet; green, isolated bridge; cyan, turn; white, random coil.

Figure 10

Evolution of secondary structures in explicit water of (A) p53: nucleation point (B) p53: ionic interaction (C) p63: nucleation point and (D) p73: nucleation point; Colour code: purple, α-helix; red, π-helix; yellow, β-sheet; green, isolated bridge; cyan, turn; white, random coil.