# Membrane-Mediated Interactions of the Bcl-xL/Bid/Bax Triad of Apoptotic Regulators

> **NIH NIH R01** · UNIVERSITY OF KANSAS MEDICAL CENTER · 2020 · $449,309

## Abstract

Summary
 Apoptosis is crucial for proper development and function of cell populations in tissues, and its dys-
regulation is of major relevance for degenerative diseases and cancer. The critical step in triggering
apoptosis is the permeabilization of the mitochondrial outer membrane (MOMP). This process is tightly
regulated by the Bcl-2 family of proteins, which is subdivided into pro-apoptotic (e.g., Bax), anti-apoptotic
(e.g., Bcl-xL), and BH3-only regulator proteins (e.g., Bid). Despite recent advances in the characterization
of Bcl-2 proteins, the field lacks a mechanistic understanding of the protein–protein and protein–lipid
interactions that mediate MOMP. Such knowledge would be essential for setting the stage for the future
development of therapeutic strategies aimed at either suppressing or activating apoptosis. The proposed
project is aimed at deciphering the pathways of membrane insertion and refolding of the Bax/Bid/Bcl-xL
regulatory triad and characterizing their membrane-modulated interactions within the framework of the
Embedded Together model of apoptotic regulation by Bcl-2 proteins. We will draw from our experience with
other membrane-inserting proteins, including the diphtheria toxin translocation domain, which has structural
similarities to Bcl-2 proteins. Site-specific labeling in combination with a battery of fluorescence (including
various types of steady-state and lifetime quenching and FRET) and electron paramagnetic resonance
approaches (DEER, O2/NiEDDA accessibility), complemented by Molecular Dynamics computer
simulations, will be utilized to obtain structural, dynamic and thermodynamic information necessary for
deciphering the mechanism of physiological function. Our preliminary data indicate that conformational
switching and activation Bid/Bax/Bcl-xL regulatory triad is modulated by the electrostatic and mechanical
properties of the lipid bilayer. We will pursue the following specific aims: (1) Characterize membrane-
dependent conformational switching in anti-apoptotic Bcl-xL, (3) Characterize lipid-dependent membrane
recruitment of the Bid/Bax/Bcl-xL regulatory triad, (3) Characterize the membrane insertion pathway of pore-
forming Bax and its disruption by Bcl-xL. By gaining new insights into molecular mechanisms of protein–
protein and protein–lipid interactions in the Bax/Bcl-xL/Bid regulatory triad, we expect to provide a clearer
map of the molecular pathways controlling MOMP. In addition, this study will reveal general principles of
protein conformational switching on membranes that will inform other regulatory systems of biomedical
importance.

## Key facts

- **NIH application ID:** 9858374
- **Project number:** 5R01GM126778-02
- **Recipient organization:** UNIVERSITY OF KANSAS MEDICAL CENTER
- **Principal Investigator:** ALEXEY LADOKHIN
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $449,309
- **Award type:** 5
- **Project period:** 2019-02-01 → 2023-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9858374

## Citation

> US National Institutes of Health, RePORTER application 9858374, Membrane-Mediated Interactions of the Bcl-xL/Bid/Bax Triad of Apoptotic Regulators (5R01GM126778-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9858374. Licensed CC0.

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