Biology subjects

Luo, Y. L.

Publications and source records attributed to Luo, Y. L..

7 recordsLinked to original sources

bioRxiv2026

Yoda molecules agonize PIEZO2

PIEZO proteins (PIEZO1 and PIEZO2) are essential mechanosensitive channels. PIEZO1 is thought to be selectively activated by Yoda molecules (Yoda1 and Yoda2). Although a structural framework for PIEZO1 activation by Yoda1 exists, a molecular mechanism underlying this selective activation is lacking. Here, using electrophysiology and calcium imaging, we show that Yoda1 increases PIEZO2 open probability and stretch sensitivity as efficaciously as PIEZO1 but elicits weaker PIEZO2-dependent calcium entry, rationalizing why its effect on PIEZO2 has been overlooked. Both Yoda1 and its more potent Yoda2 analog slow down inactivation of PIEZO2 currents with potency similar to PIEZO1 but with lower efficacy. Using mutagenesis and molecular dynamics simulations, we further show that Yoda2s benzoic acid group forms a transient salt bridge with a conserved arginine in the Yoda binding site, providing a molecular basis for Yoda2s increased potency. Our study cautions a reevaluation of studies using these molecules to untangle biological functions mediated by PIEZO channels.

biophysics↗

bioRxiv2026

CholBindNet: Interpretable Neural Networks for Cholesterol Binding Site Prediction

Cholesterol is a key modulator of membrane protein structure and function, yet predicting cholesterol binding sites remains challenging due to its undrug-like physicochemical properties. Here, we curated more than 800 high-resolution transmembrane protein structures containing cholesterol and developed an interpretable, atom-based deep-learning framework, CholBindNet, comprising four model architectures: a 3D convolutional neural network, a graph neural network, a graph attention network, and a graph convolutional network. A Positive-Unlabeled (PU) training strategy was employed to address the scarcity of true negative samples resulting from the promiscuous nature of cholesterol binding. We show that CholBindNet substantially outperforms existing deep-learning models trained on general ligand-binding datasets. The performance and generalizability of the model were further demonstrated by rapidly assessing strong, median, and weak cholesterol-binding sites in the PIEZO2 ion channel in excellent agreement with computationally expensive all-atom molecular dynamics (MD) simulations. Additionally, strong model interpretability was achieved for CholBindNet through atom-level feature encoding, Grad-CAM visualization, and attention-based scoring analysis. Overall, CholBindNet provides an efficient and scalable approach for predicting cholesterol binding sites on membrane proteins, achieving performance comparable to MD simulations while offering mechanistic biophysical insights beyond amino-acid sequence. This work hence lays the foundation for future development of deep-learning models targeting membrane protein drug-binding sites and cholesterol-modulated therapeutics. Significance StatementDeep-learning models for ligand-binding prediction have advanced rapidly, yet those trained on soluble proteins perform poorly for membrane proteins, particularly for cholesterol binding. We introduce CholBindNet, a set of neural network models specifically designed to identify cholesterol-binding sites in transmembrane proteins. CholBindNet substantially outperforms existing deep-learning approaches and accurately ranks strong, intermediate, and weak cholesterol-binding sites in close agreement with computationally intensive all-atom molecular dynamics simulations. This work provides a practical and scalable alternative to long-timescale simulations for studying cholesterol-protein interactions. The curated cholesterol benchmark and open-source models will enable broader adoption of deep learning for investigating lipid regulation of membrane proteins and for guiding the design of drugs targeting membrane-embedded binding sites.

biophysics↗

bioRxiv2025

A two-step clockwork mechanism opens a proteo-lipidic pore in PIEZO2 channels

Mechanosensitive PIEZO channels are thought to open their pore through tension-induced flattening of large transmembrane arm domains. Yet, the structural basis of this activation remains unclear. Here, we uncover the conformational coupling between arm flattening and pore opening in PIEZO2 by capturing protein motions across length scales using hybrid-resolution molecular dynamics simulations. Sampling multiple microsecond-long trajectories under physiological activation tension show that arm flattening correlates with anticlockwise rotation of the pore domain and with clockwise twisting of inner pore helices, enabling dilation and hydration of a transmembrane pore gate. These clockwork motions enable PIEZO2 to populate two open states with distinct conductance depending on applied membrane tension, in agreement with single-channel electrophysiology. Pore opening is accompanied by the separation of pore helices, creating interhelical gaps which become filled with lipids, resulting in the fully conducting pore being walled by both lipids and protein. The fully open PIEZO2 state recapitulates minimal pore size, conductance, ion selectivity, and outward rectification of chloride currents observed electrophysiologically. This work reveals how tension-induced large-scale rearrangements of the PIEZO2 arms funnel into subtle and dynamic gating motions, providing invaluable structural insights for future structure-function and drug discovery studies.

biophysics↗

bioRxiv2024

Dissecting current rectification through asymmetric nanopores

Rectification, the tendency of bidirectional ionic conductors to favor ion flow in a specific direction, is an intrinsic property of many ion channels and synthetic nanopores. Despite its frequent occurrence in ion channels and its phenomenological explanation using Eyrings rate theory, a quantitative relationship between the rectified current and the underlying ion-specific and voltage-dependent free energy profile has been lacking. In this study, we designed nanopores in which potassium and chloride current rectification can be manipulated by altering the electrostatic pore polarity. Using molecular dynamics-based free energy simulations, we quantified voltage-dependent changes of free energy barriers in six ion-nanopore systems. Our results illustrate how the energy barriers for inward and outward fluxes become unequal in the presence of an electromotive driving force, leading to varying degrees of rectification for cation and anion currents. This work establishes a direct link between equilibrium potential of mean force and current rectification rate and demonstrates that rectification caused by energy barrier asymmetry depends on the nature of the permeating ion, can be tuned by pore polarity, does not require ion binding sites, conformational flexibility, or specific pore geometry, and, as such, may be widespread among ion channels. Statement of significanceIn many ion channels, ions flow faster in one direction than the other, even under equal magnitude of driving forces. This phenomenon, known as rectification, often arises as an intrinsic biophysical property of the channel pore and has significant physiological implications. Our molecular dynamics simulations provide a unified framework to dissect the free energy profiles that underpin current rectification. This approach has broad applications in the field of ion channel biology and material design.

biophysics↗

bioRxiv2024

Optimizing Coarse-Grained Models for Large-Scale Membrane Protein Simulation

Coarse-grained (CG) models have been developed for studying membrane proteins at physiologically relevant scales. Such methods, including popular CG lipid models, exhibit stability and efficiency at moderate scales, but they can become impractical or even unusable beyond a critical size due to various technical issues. Here, we report that these scale-dependent issues can arise from progressively slower relaxation dynamics and become confounded by unforeseen instabilities observed only at larger scales. To address these issues, we systemically optimized a 4-site solvent-free CG lipid model that is suitable for conducting micron-scale molecular dynamics simulations of membrane proteins under various membrane properties. We applied this lipid model to explore the long-range membrane deformation induced by a large mechanosensitive ion channel, PIEZO. We show that the optimized CG models are powerful in elucidating the structural and dynamic interplay between PIEZO and the membrane. Furthermore, we anticipate that our methodological insights can prove useful for resolving issues stemming from scale-dependent limitations of similar CG methodologies.

biophysics↗

bioRxiv2024

Large-pore connexin hemichannels function as molecule transporters independently of ion conduction

Connexin hemichannels were identified as the first members of the eukaryotic large-pore channel family that mediate permeation of both atomic ions and small molecules between the intracellular and extracellular environments. The conventional view is that their pore is a large passive conduit through which both ions and molecules diffuse in a similar manner. In stark contrast to this notion, we demonstrate that the permeation of ions and of molecules in connexin hemichannels can be uncoupled and differentially regulated. We find that human connexin mutations that produce pathologies and were previously thought to be loss-of-function mutations due to the lack of ionic currents are still capable of mediating the passive transport of molecules with kinetics close to those of wild-type channels. This molecular transport displays saturability in the micromolar range, selectivity, and competitive inhibition, properties that are tuned by specific interactions between the permeating molecules and the N-terminal domain that lies within the pore -- a general feature of large-pore channels. We propose that connexin hemichannels and, likely, other large-pore channels, are hybrid channel/transporter-like proteins that might switch between these two modes to promote selective ion conduction or autocrine/paracrine molecular signaling in health and disease processes.

biophysics↗

bioRxiv2020

A Piezo1 open state reveals a multi-fenestrated ion permeation pathway

Force-sensing Piezo channels are essential to many aspects of vertebrate physiology. Activation of Piezo1 is facilitated by the presence of negative membrane lipids in the inner leaflet, such as phosphatidylinositol-4,5-bisphosphate (PIP2). Here, to study how Piezo1 opens, we performed molecular dynamics simulations of Piezo1 in membranes flattened by the periodic boundary effect and with or without PIP2 lipids. The Piezo1 pore spontaneously opens in the asymmetrical bilayer but not in the symmetric membrane or when PIP2 lipids are neutralized. Electrophysiological characterization of putative PIP2-interacting Piezo1 residues suggests the contribution of multiple PIP2 binding sites. Our Piezo1 open state recapitulates ionic selectivity, unitary conductance and mutant phenotypes obtained from numerous experimental studies. Tracking ion diffusion through the open pore reveals the presence of intracellular and extracellular fenestrations, delineating a multi-fenestrated permeation pathway. This open state sheds light on the mechanisms of lipid modulation, permeation, and selectivity in a Piezo channel.

biophysics↗