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Yeast As a Chassis for Developing Functional Assays to Study Human P53
Published on: August 4, 2019
Modulation of p53 Transactivation Domain Conformations by Ligand Binding and Cancer-Associated Mutations
1Department of Chemistry, University of Massachusetts Amherst, Amherst, MA 01003, USA.
Abstract:
Intrinsically disordered proteins (IDPs) are important functional proteins, and their deregulation are linked to numerous human diseases including cancers. Understanding how disease-associated mutations or drug molecules can perturb the sequence-disordered ensemble-function-disease relationship of IDPs remains challenging, because it requires detailed characterization of the heterogeneous structural ensembles of IDPs. In this work, we combine the latest atomistic force field a99SB-disp, enhanced sampling technique replica exchange with solute tempering, and GPU-accelerated molecular dynamics simulations to investigate how four cancer-associated mutations, K24N, N29K/N30D, D49Y, and W53G, and binding of an anti-cancer molecule, epigallocatechin gallate (EGCG), modulate the disordered ensemble of the transactivation domain (TAD) of tumor suppressor p53. Through extensive sampling, in excess of 1.0 μs per replica, well-converged structural ensembles of wild-type and mutant p53-TAD as well as WT p53-TAD in the presence of EGCG were generated. The results reveal that mutants could induce local structural changes and affect secondary structural properties. Interestingly, both EGCG binding and N29K/N30D could also induce long-range structural reorganizations and lead to more compact structures that could shield key binding sites of p53-TAD regulators. Further analysis reveals that the effects of EGCG binding are mainly achieved through nonspecific interactions. These observations are generally consistent with on-going NMR studies and binding assays. Our studies suggest that induced conformational collapse of IDPs may be a general mechanism for shielding functional sites, thus inhibiting recognition of their targets. The current study also demonstrates that atomistic simulations provide a viable approach for studying the sequence-disordered ensemble-function-disease relationships of IDPs and developing new drug design strategies targeting regulatory IDPs.
Insights
Cancer-linked mutations and EGCG binding alter the structure of p53 transactivation domain (TAD). These changes can shield crucial binding sites, suggesting a new mechanism for disease and drug development targeting intrinsically disordered proteins (IDPs).
Area of Science:
- Biochemistry and Molecular Biology
- Computational Biology
- Structural Biology
Background:
- Intrinsically disordered proteins (IDPs) lack stable structures but are crucial for cellular functions, and their dysregulation is implicated in diseases like cancer.
- Understanding how mutations or drugs affect IDP structure and function is vital for disease research and therapeutic development.
- Characterizing the dynamic, heterogeneous structural ensembles of IDPs presents a significant scientific challenge.
Purpose of the Study:
- To investigate the impact of cancer-associated mutations and anti-cancer drug binding on the structural ensemble of the p53 transactivation domain (TAD).
- To explore the sequence-disordered ensemble-function-disease relationships of IDPs using advanced computational methods.
- To elucidate the molecular mechanisms by which drug molecules modulate IDP structure and potentially inhibit function.
Main Methods:
- Utilized GPU-accelerated molecular dynamics simulations with the a99SB-disp force field and replica exchange with solute tempering (REST2) enhanced sampling.
- Generated well-converged structural ensembles for wild-type and mutant p53-TAD, and for wild-type p53-TAD in the presence of epigallocatechin gallate (EGCG).
- Performed extensive sampling exceeding 1.0 μs per replica to capture the heterogeneous nature of IDP ensembles.
Main Results:
- Identified that cancer-associated mutations (K24N, N29K/N30D, D49Y, W53G) induce local structural changes and affect secondary structure properties of p53-TAD.
- Observed that both EGCG binding and the N29K/N30D mutation promote long-range structural reorganizations, leading to more compact p53-TAD structures.
- Found that EGCG binding primarily acts through non-specific interactions, potentially shielding key regulatory binding sites on p53-TAD.
Conclusions:
- Induced conformational collapse of IDPs may represent a general mechanism for shielding functional sites, thereby inhibiting target recognition.
- Atomistic simulations offer a powerful and viable approach for dissecting IDP ensemble-function-disease relationships.
- This study provides insights for developing novel drug design strategies targeting regulatory IDPs in cancer therapy.
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