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Investigating DNA Binding and Conformational Variation in Temperature Sensitive p53 Cancer Mutants Using QM-MM
Shruti Koulgi1, Archana Achalere1, Uddhavesh Sonavane1
1Bioinformatics Group, Center for Development of Advanced Computing (C-DAC), S.P.Pune University Campus, Pune, India.
Abstract:
The tp53 gene is found to be mutated in 50% of all the cancers. The p53 protein, a product of tp53 gene, is a multi-domain protein. It consists of a core DNA binding domain (DBD) which is responsible for its binding and transcription of downstream target genes. The mutations in p53 protein are responsible for creating cancerous conditions and are found to be occurring at a high frequency in the DBD region of p53. Some of these mutations are also known to be temperature sensitive (ts) in nature. They are known to exhibit partial or strong binding with DNA in the temperature range (298-306 K). Whereas, at 310 K and above they show complete loss in binding. We have analyzed the changes in binding and conformational behavior at 300 K and 310 K for three of the ts-mutants viz., V143A, R249S and R175H. QM-MM simulations have been performed on the wild type and the above mentioned ts-mutants for 30 ns each. The optimal estimate of free energy of binding for a particular number of interface hydrogen bonds was calculated using the maximum likelihood method as described by Chodera et. al (2007). This parameter has been observed to be able to mimic the binding affinity of the p53 ts-mutants at 300 K and 310 K. Thus the correlation between MM-GBSA free energy of binding and hydrogen bonds formed by the interface residues between p53 and DNA has revealed the temperature dependent nature of these mutants. The role of main chain dihedrals was obtained by performing dihedral principal component analysis (PCA). This analysis, suggests that the conformational variations in the main chain dihedrals (ϕ and ψ) of the p53 ts-mutants may have caused reduction in the overall stability of the protein. The solvent exposure of the side chains of the interface residues were found to hamper the binding of the p53 to the DNA. Solvent Accessible Surface Area (SASA) also proved to be a crucial property in distinguishing the conformers obtained at 300 K and 310 K for the three ts-mutants from the wild type at 300 K.
Insights
Temperature-sensitive p53 mutants show altered DNA binding and stability at different temperatures. Computational analysis reveals how these mutations impact protein conformation and binding affinity, crucial for cancer development.
Area of Science:
- Molecular Biology
- Computational Biophysics
- Cancer Genetics
Background:
- The tp53 gene is frequently mutated in 50% of human cancers.
- p53 protein mutations, particularly in the DNA binding domain (DBD), are linked to cancer development.
- Some p53 mutations are temperature-sensitive (ts), affecting DNA binding at physiological temperatures.
Purpose of the Study:
- To analyze the binding and conformational changes of three temperature-sensitive p53 mutants (V143A, R249S, R175H) at 300 K and 310 K.
- To understand the molecular mechanisms underlying the temperature-dependent loss of p53-DNA binding affinity.
- To correlate computational findings with the role of these mutations in cancer.
Main Methods:
- Performed 30 ns QM-MM simulations on wild-type and three ts-p53 mutants.
- Calculated free energy of binding using the maximum likelihood method based on interface hydrogen bonds.
- Utilized dihedral principal component analysis (PCA) to study main chain dihedral variations.
- Analyzed Solvent Accessible Surface Area (SASA) to assess side chain solvent exposure.
Main Results:
- A correlation between MM-GBSA free energy of binding and interface hydrogen bonds revealed the temperature-dependent nature of p53 ts-mutants.
- Conformational variations in main chain dihedrals (ϕ and ψ) of ts-mutants may reduce protein stability.
- Increased solvent exposure of interface residues' side chains hampers p53-DNA binding.
- SASA effectively distinguished conformers of ts-mutants at different temperatures from wild-type p53.
Conclusions:
- The study elucidates the temperature-dependent binding and conformational dynamics of critical p53 mutants.
- Computational methods accurately mimic the binding affinity changes observed in ts-p53 mutants.
- Findings provide insights into how p53 mutations contribute to cancer by altering DNA binding and protein stability.
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