Comprehensive Identification of Deleterious TP53 Missense VUS Variants Based on Their Impact on TP53 Structural Stability
Abstract
:1. Introduction
2. Results
2.1. Construction of Mutant Protein Structures
2.2. Identification of Deleterious Variants
2.3. Features of Deleterious VUS
- Deviation distribution. The range of deviation was 3.158–3.878 (23.5–48.3%) (Figure 3). This range was similar to that of known pathogenic variants, except for the top 3—R273C, Y220C, R175H (49.6, 47.4, 45.6%)—as they were highly destructive for the TP53 structure (Figure 2). For benign variants, 6 out of 23 benign variants were above the deleterious structure limits (>23.5%). Here, the Q165K had a structure deviation of 35.7% and was an outlier within the benign variants. The variants below the cutoff line were classified as “unknown”, as certain pathogenic variants may have minimal impact on structure stability [19]; therefore, their overall structure scaffolds can be comparable to benign variants. Overall, the results justified the use of the deviation from known pathogenic variants as the reference to classify the missense VUS.
- Spatial change of the substituted residues. The Ramachandran plot (RP) showed the spatial differences of the substituted residues from the wildtype residues, and the root-mean-square-deviation (RMSD) plot also showed the altered position of the substituted residues from the wildtype residues in the global TP53 structure (Figure 4A–C). For example, in S99F, F in RP showed its torsional angle in reflecting the rigidity of the fluctuation, and F in RMSD also showed its larger fluctuation, revealing its instability in the local environment; in G154R, R in RP showed its torsional angle fluctuation deviated from the wildtype G, and R in RMSD showed its large fluctuation, reflecting its instability in its local environment; in H214P, P in RP showed its torsional angle substantially fluctuated from the wildtype residue H, and the lower RMSD showed its high stability in TP53.
- Distribution in TP53 secondary structure. The deleterious VUS variants were distributed across the entire DNA binding domain, of which 44 were in the regions with few known pathogenic variants, including entire β sheets, loop 1 and loop 2, and all linkers; for the regions overlapped with the known pathogenic variants, the deleterious VUS variants were distributed more widely than the known pathogenic variants (Figure 5). The results indicate that RPMDS provides high sensitivity to detect deleterious missense VUS variants.
- Impact on TP53 local structure. The Ramachandran density plots showed that the deleterious VUS variants caused more local structural change, whereas known pathogenic variants caused more global structural change (Figure 6A). Taking Y107D, M169V, R249S, T253N, and I255S as examples (Figure 6B): in Y107D, the change of residue caused greater flexibility in the β strand (residues 108–114 (β1) and 204–208 (β6)); this was reflected by the diminishing peak in the Ramachandran density plot: an α helix loop was formed between residues 165–172, and this was not observed in WT; in M169V, the flexible structure was reflected by the diminished peaks at P-II and the β sheet region in the Ramachandran density plot. The extra α-helix bend was formed at residues 117–121 and 168–170, and α-helix 1 (177–181) was more structurally stable than the WT α-helix 1 (H1); with its high deviation of 3.770 (43.4%), R249S misfolded the TP53 structure globally, and the Ramachandran plot revealed a spike at the P-II region, while the β sheet region was diminished; in T253N, greater flexibility of β strand S4 and shortened β strand S8 (residues 156–161 and 230–236, respectively) reflected the diminished peak of the Ramachandran density plot; the residues 165–172 showed a stable structural formation, and this was not detected in the WT; in I255S, the Ramachandran density plot revealed the dissipated peak at the β sheet and P-II spiral regions; higher flexibility was present at the linker residues (244–246) and α-helix 2 (H2) (285–286), and S10 β strand (271–274) was extended by 2 residues.
2.4. Comparison with Other In Silico Methods
3. Discussion
4. Materials and Methods
4.1. Source of Missense VUS Variants
4.2. Molecular Dynamics Simulation (MDS)
4.3. Ramachandran Plot (RP)
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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(A) Comparison with Updated ClinVar Classification | ||||
Variant | ClinVar Classification | RPMDS Classification | ||
Nucleotide | Amino Acid | Original | New | |
c.706T>G | Y236D | VUS | Pathogenic | Deleterious |
c.695T>A | I232N | VUS | Pathogenic | Deleterious |
c.794T>A | L265Q | VUS | Pathogenic | Deleterious |
c.413C>T | A138V | VUS | Conflict | Deleterious |
c.422G>T | C141F | VUS | Conflict | Deleterious |
c.434T>C | L145P | VUS | Conflict | Deleterious |
c.526T>A | C176S | VUS | Conflict | Deleterious |
c.556G>A | D186N | VUS | Conflict | Deleterious |
c.581T>G | L194R | VUS | Conflict | Deleterious |
c.626G>A | R209K | VUS | Conflict | Deleterious |
c.814G>A | V272M | VUS | Conflict | Deleterious |
c.931A>C | N311H | VUS | Conflict | Deleterious |
c.431A>T | Q144L | VUS | Conflict | Undefined |
c.452C>G | P151R | VUS | Conflict | Undefined |
c.658T>C | Y220H | VUS | Conflict | Undefined |
c.730G>T | G244C | VUS | Conflict | Undefined |
c.928A>G | N310D | VUS | Conflict | Undefined |
(B) Comparison with Different In Silico Methods in Classifying 340 Missense VUS | ||||
Methods | Classification | |||
Deleterious * | Rate (%) | Undefined ** | Rate (%) | |
RPMDS | 193 | 56.8 | 147 | 43.2 |
Polyphen2_HVAR_pred | 202 | 59.4 | 138 | 40.6 |
Polyphen2_HDIV_pred | 217 | 62.9 | 123 | 37.1 |
PROVEAN_pred | 233 | 68.5 | 107 | 31.5 |
LRT_pred | 242 | 71.2 | 98 | 28.8 |
SIFT_pred | 268 | 78.8 | 72 | 21.2 |
MutationTaster_pred | 287 | 84.4 | 53 | 15.6 |
M-CAP_pred | 335 | 98.5 | 5 | 1.5 |
FATHMM_pred | 340 | 100 | 0 | 0 |
MetaSVM_pred | 340 | 100 | 0 | 0 |
MetaLR_pred | 340 | 100 | 0 | 0 |
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Tam, B.; Sinha, S.; Qin, Z.; Wang, S.M. Comprehensive Identification of Deleterious TP53 Missense VUS Variants Based on Their Impact on TP53 Structural Stability. Int. J. Mol. Sci. 2021, 22, 11345. https://doi.org/10.3390/ijms222111345
Tam B, Sinha S, Qin Z, Wang SM. Comprehensive Identification of Deleterious TP53 Missense VUS Variants Based on Their Impact on TP53 Structural Stability. International Journal of Molecular Sciences. 2021; 22(21):11345. https://doi.org/10.3390/ijms222111345
Chicago/Turabian StyleTam, Benjamin, Siddharth Sinha, Zixin Qin, and San Ming Wang. 2021. "Comprehensive Identification of Deleterious TP53 Missense VUS Variants Based on Their Impact on TP53 Structural Stability" International Journal of Molecular Sciences 22, no. 21: 11345. https://doi.org/10.3390/ijms222111345