Related Experiment Video
Updated: Jun 14, 2025
Fluorescence-Based Measurements of Phosphatidylserine/Phosphatidylinositol 4-Phosphate Exchange Between Membranes
Published on: March 14, 2021
Free energy landscape of the PI3Kα C-terminal activation
Danai Maria Kotzampasi1,2, Michail Papadourakis1, John E Burke3,4
1Biomedical Research Foundation, Academy of Athens, Athens 11527, Greece.
Abstract:
The gene PIK3CA, encoding the catalytic subunit p110α of PI3Kα, is the second most frequently mutated gene in cancer, with the highest frequency oncogenic mutants occurring in the C-terminus of the kinase domain. The C-terminus has a dual function in regulating the kinase, playing a putative auto-inhibitory role for kinase activity and being absolutely essential for binding to the cell membrane. However, the molecular mechanisms by which these C-terminal oncogenic mutations cause PI3Kα overactivation remain unclear. To understand how a spectrum of C-terminal mutations of PI3Kα alter kinase activity compared to the WT, we perform unbiased and biased Molecular Dynamics simulations of several C-terminal mutants and report the free energy landscapes for the C-terminal "closed-to-open" transition in the WT, H1047R, G1049R, M1043L and N1068KLKR mutants. Results are consistent with HDX-MS experimental data and provide a molecular explanation why H1047R and G1049R reorient the C-terminus with a different mechanism compared to the WT and M1043L and N1068KLKR mutants. Moreover, we show that in the H1047R mutant, the cavity, where the allosteric ligands STX-478 and RLY-2608 bind, is more accessible contrary to the WT. This study provides insights into the molecular mechanisms underlying activation of oncogenic PI3Kα by C-terminal mutations and represents a valuable resource for continued efforts in the development of mutant selective inhibitors as therapeutics.
Insights
The PIK3CA gene
Area of Science:
- Biochemistry
- Molecular Biology
- Cancer Research
Background:
- The PIK3CA gene encodes the p110α catalytic subunit of PI3Kα, a key enzyme in cell signaling.
- Mutations in PIK3CA are common in cancer, particularly in the C-terminus of the kinase domain, leading to overactivation.
- The precise molecular mechanisms by which C-terminal PIK3CA mutations drive oncogenesis are not fully understood.
Purpose of the Study:
- To elucidate the molecular mechanisms of PI3Kα overactivation caused by C-terminal mutations.
- To compare the kinase activity and conformational changes of wild-type (WT) PI3Kα with various C-terminal mutants.
- To provide a molecular basis for the development of targeted cancer therapies.
Main Methods:
- Unbiased and biased Molecular Dynamics (MD) simulations were employed to study PI3Kα C-terminal mutants.
- Free energy landscapes of the C-terminal "closed-to-open" transition were calculated for WT and mutant forms.
- Results were validated against experimental Hydrogen-Deuterium Exchange Mass Spectrometry (HDX-MS) data.
Main Results:
- MD simulations revealed distinct mechanisms of C-terminal reorientation for H1047R and G1049R mutants compared to WT, M1043L, and N1068KLKR mutants.
- The H1047R mutation increases the accessibility of the allosteric ligand-binding pocket.
- Simulation results align with experimental HDX-MS data, supporting the proposed mechanisms.
Conclusions:
- This study offers molecular insights into how C-terminal PIK3CA mutations activate PI3Kα.
- The findings clarify the differential effects of specific mutations on enzyme conformation and activity.
- The research serves as a foundation for designing mutant-specific PI3Kα inhibitors for cancer treatment.
Related Concept Videos
01:22PI3K/mTOR/AKT Signaling Pathway
01:20The JAK-STAT Signaling Pathway
01:11IP3/DAG Signaling Pathway
01:22Amplifying Signals via Enzymatic Cascade
01:42Phosphoinositides and PIPs
Different phosphoinositides are synthesized and recruited on the cytosolic face of the plasma membrane. The localization of specific phosphoinositides concentrated in separate membrane...
01:19Interactions Between Signaling Pathways
Convergence and divergence, and cross-talk between signaling pathways
Two distinct signaling pathways can converge on a single functional unit, which may either be a single protein or a complex of proteins. The response is either functionally distinct or synergistic between the two pathways but different from the response...