Phosphoinositides protonation states dictate AKT1 binding with membrane
- 1Indian Institute of Science
Abstract
Phosphoinositides are key regulators of membrane-associated signaling, yet their electrostatic properties are commonly represented by a single nominal charge in molecular simulations and structural models. Using the essential protein kinase AKT1 as a model system, we show that phosphoinositide protonation microstates, both the degree and positional placement of protons, govern peripheral membrane-protein recognition. By integrating quantum-mechanics-derived headgroup parameters, microsecond-scale all-atom molecular dynamics, and umbrella-sampling free-energy calculations with the available solid-state NMR-derived lipid populations via Bayesian/Maximum Entropy reweighting, we account explicitly for the thermodynamic ensemble of phosphoinositide proto-nation states. This population-weighted free-energy framework provides a quantitative mechanistic rationale for the strict specificity of wild-type AKT1 toward PI(3, 4, 5)P3 and elucidates how the oncogenic sentry mutation (E17K) alters this specificity to enable high-affinity PIP2 binding. Furthermore, Proximity-based Labeling of Membrane Associated Proteins (PLiMAP) assays experimentally confirm that AKT1 pleckstrin homology domain binding to distinct phosphoinositide species is differentially sensitive to pH. Together, our findings establish a direct connection between phosphoinositide protonation equilibria, binding energetics, and AKT1 lipid specificity. Beyond AKT1, these results demonstrate that the dynamic protonation microstates of anionic phospholipids act as a functional regulatory layer governing peripheral membrane-protein recruitment across diverse cellular microenvironments.
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BibTeX
@misc{jha2026phosphoinositides,
title = {{Phosphoinositides protonation states dictate AKT1 binding with membrane}},
author = {K. Jha and M. Sarkar and K. Baratam and J. Nagesh and T. Pucadyil and A. Srivastava},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.05.749365},
url = {https://www.biorxiv.org/content/10.64898/2026.09.05.749365v1},
note = {CC BY-NC},
}
RIS
TY - UNPB
TI - Phosphoinositides protonation states dictate AKT1 binding with membrane
AU - K. Jha
AU - M. Sarkar
AU - K. Baratam
AU - J. Nagesh
AU - T. Pucadyil
AU - A. Srivastava
PY - 2026
DA - 2026/09/05
PB - bioRxiv
DO - 10.64898/2026.09.05.749365
UR - https://www.biorxiv.org/content/10.64898/2026.09.05.749365v1
AB - Phosphoinositides are key regulators of membrane-associated signaling, yet their electrostatic properties are commonly represented by a single nominal charge in molecular simulations and structural models. Using the essential protein kinase AKT1 as a model system, we show that phosphoinositide protonation microstates, both the degree and positional placement of protons, govern peripheral membrane-protein recognition. By integrating quantum-mechanics-derived headgroup parameters, microsecond-scale all-atom molecular dynamics, and umbrella-sampling free-energy calculations with the available solid-state NMR-derived lipid populations via Bayesian/Maximum Entropy reweighting, we account explicitly for the thermodynamic ensemble of phosphoinositide proto-nation states. This population-weighted free-energy framework provides a quantitative mechanistic rationale for the strict specificity of wild-type AKT1 toward PI(3, 4, 5)P3 and elucidates how the oncogenic sentry mutation (E17K) alters this specificity to enable high-affinity PIP2 binding. Furthermore, Proximity-based Labeling of Membrane Associated Proteins (PLiMAP) assays experimentally confirm that AKT1 pleckstrin homology domain binding to distinct phosphoinositide species is differentially sensitive to pH. Together, our findings establish a direct connection between phosphoinositide protonation equilibria, binding energetics, and AKT1 lipid specificity. Beyond AKT1, these results demonstrate that the dynamic protonation microstates of anionic phospholipids act as a functional regulatory layer governing peripheral membrane-protein recruitment across diverse cellular microenvironments.
ER -