Tumorigenic p53 mutants undergo common structural disruptions including conversion to α-sheet structure

Abstract

The p53 protein is a commonly studied cancer target because of its role in tumor suppression. Unfortunately, it is susceptible to mutation-associated loss of function; approximately 50% of cancers are associated with mutations to p53, the majority of which are located in the central DNA-binding domain. Here, we report molecular dynamics simulations of wild-type (WT) p53 and 20 different mutants, including a stabilized pseudo-WT mutant. Our findings indicate that p53 mutants tend to exacerbate latent structural-disruption tendencies, or vulnerabilities, already present in the WT protein, suggesting that it may be possible to develop cancer therapies by targeting a relatively small set of structural-disruption motifs rather than a multitude of effects specific to each mutant. In addition, α-sheet secondary structure formed in almost all of the proteins. α-Sheet has been hypothesized and recently demonstrated to play a role in amyloidogenesis, and its presence in the reported p53 simulations coincides with the recent re-consideration of cancer as an amyloid disease.

Keywords: aggregation; cancer; molecular dynamics; visual analytics; α-sheet.

FIGURE 1

(a) p53 crystal structure (Protein Data Bank [PDB]: 2OCJ) labeled with secondary‐structure. Zinc is depicted with a blue ball. (b) Schematic overview of the p53 secondary structures in (a). (c) p53 crystal structure overlaid with the mutations presented here

FIGURE 2

Crystal structure of stabilized pseudo‐wild type (WT; Protein Data Bank [PDB]: 1UOL) overlaid with the 81 contacts whose occupancy differed by at least 50% from WT. The four quad‐stabilizing mutation sites (M133L, V203A, N239Y, N268D) are shown as spheres

FIGURE 3

Rotated views of the β‐sheet solvent exposure of the 2OCJ crystal structure. β‐Sheets are colored and the remainder of the structures are gray. (Top) Solvent‐accessible surface area. (Bottom) Cartoon view

FIGURE 4

p53 structures highlighting disrupted loop–sheet–helix region contacts. Cα atoms of K120 and R280 are colored cyan, stabilizing mutation site M133 is colored yellow. (a) p53 crystal structure (2OCJ). K120/R280 distance is 6.7 Å. (b) p53 wild‐type (WT) MD structure at 80 ns. K120/R280 distance is 20 Å. (c) p53 crystal structure overlaid with contacts common to all 17 L1/H2‐separated conformations. Green lines indicate contacts that were present in the separated conformations but not in the crystal‐like WT conformation. Magenta lines indicate contacts that were not present in the separated conformations and were present in the crystal‐like WT conformation. Table S3 specifies which contacts were present in the separated and nonseparated conformations

FIGURE 5

p53 structures highlighting disrupted L2 and S5 region contacts. Cα atoms of D186 and G199 are colored cyan, stabilizing mutation site V203 is colored yellow. (a) p53 crystal structure (2OCJ). D186/G199 distance is 10.6 Å. (b) p53 R248Q mutant MD structure at 80 ns. D186/G199 distance is 24.6 Å. (c) p53 crystal structure overlaid with contacts common to all 14 L2/S5‐separated conformations. Green lines indicate contacts that were present in the separated conformations but not in the crystal‐like wild‐type (WT) conformation. Magenta lines indicate contacts that were not present in the separated conformations and were present in the crystal‐like WT conformation. Table S4 specifies which contacts were present in the separated and nonseparated conformations

FIGURE 6

Wild‐type (WT) apo simulation displaying α‐sheet conformation. (a) Strands S1 and S3 displaying β‐sheet conformation at 0 ns. (b) Strands S1 and S3 displaying α‐sheet conformation at 75 ns

FIGURE 7

Proposed G245S α‐sheet formation model. Arrows indicate new motion and fade over time. (a) 0 and (b) 31 ns. N‐terminal Q104:OE1 orients to F109:O, causing it to rotate and face W146:O. Main chain hydrogens of F109 and R110 now face Q104:OE1. (c) 45 ns. W146 rotates away from F109. (d) 60 ns. L111:O rotates inward and S1 and S3 slide past each other, finding a stable conformation one residue offset from previous conformation. (e) 85 ns. Q144 and V143 rotate and move toward F113. Note that Q104:OE1, F109:H and R110:H, once in position, remain stable for the entire process. (f) Electrostatic surface showing solvent‐accessible charge‐separation across the α‐sheet

FIGURE 8

Wild‐type S6/S7 α‐sheet. S6/S7 α‐sheet was present for 91% of the time. Structure shown at 80 ns

FIGURE 9

Novel N‐terminal α‐sheet. F134L mutant at 75.4 ns. α‐Sheet formed for 1% of the time. G105 and G266 α conformations were correlated with R = .91

FIGURE 10

p53 and transthyretin (TTR). p53 S1 aligns with TTR Strand G and p53 S3 aligns with TTR Strand A. Aligned α‐strand residues are colored green. (a) p53 crystal (Protein Data Bank [PDB]: 2OCJ). (b) TTR crystal (PDB: 1TTA)