Pore-forming proteins serve as critical biological components across multiple life forms, ranging from bacteria to humans. In humans, these proteins contribute significantly to immune defense mechanisms, creating channels that enable the passage of ions and molecules through cell membranes. In certain bacteria, pore-forming proteins function as potent toxins that disrupt cellular integrity by punching holes in membranes. The inherent ability of these biological pores to regulate molecular transport has also positioned them as invaluable assets in the rapidly evolving field of biotechnology, particularly in DNA sequencing and molecular sensing applications.
Despite their broad functional importance, the behavior of biological nanopores remains partly enigmatic, especially concerning the mechanisms driving ion transport through them. Ion flow in these nanopores exhibits complex patterns that scientists have not yet entirely deciphered. Two phenomena, in particular, have posed significant challenges: rectification and gating. Rectification describes the scenario where ion transport varies depending on the polarity of the applied voltage, effectively making the ion flow asymmetric. Gating, on the other hand, refers to abrupt reductions or stoppages in ion flow, potentially compromising the stability and reliability of nanopore-based sensing technologies.
A major breakthrough addressing these enigmas has emerged from a collaborative research team led by Matteo Dal Peraro and Aleksandra Radenovic at EPFL. Incorporating a multidisciplinary approach that blends experiments, computational modeling, and theoretical frameworks, their work meticulously unravels the fundamental principles dictating the rectification and gating behaviors in biological nanopores. This research not only sheds light on the biophysical underpinnings of these phenomena but also paves the way for enhanced design strategies in nanopore technologies.
The team centered their investigations on aerolysin, a β-barrel pore-forming protein derived from bacteria, which has found extensive use in molecular sensing due to its reliable ion channel properties. Through precision genetic engineering techniques, the researchers systematically introduced mutations to charged amino acids lining the inner surface of the nanopore. These mutations generated an extensive library of 26 unique nanopore variants, each exhibiting distinct electrical charge distributions. Comprehensive ionic current measurements through these variant nanopores under diverse voltage conditions then provided unprecedented insights into how specific charge patterns influence ion transport dynamics.
A novel aspect of the study was the use of alternating voltage signals to probe the nanopores at varying timescales. This methodological innovation enabled the researchers to temporally segregate rectification phenomena, manifesting at shorter timescales, from the more temporally extended gating events. By overlaying biophysical models with empirical data, the team constructed a robust theoretical scaffold that explains the coupling between ionic currents and nanopore structural responses—elucidating how charge localization governs the complex ion transport behaviors.
Delving into rectification, the study reveals that the distribution of electrical charges molded along the lumen of the nanopore significantly biases ion transport directionality. This intrinsic asymmetry in charge arrangement functions akin to an ionic diode or one-way valve, facilitating greater ion passage in one direction over the other. Such rectified ion flows, dictated by the electrostatic landscape, have vital implications for the sensitivity and selectivity of nanopore sensors and for the fundamental understanding of biological ion channels.
Regarding gating, the findings indicate that sustained high ionic flow can induce localized charge imbalances within the pore mouth, resulting in structural destabilization. This destabilization causes partial collapse or constriction of the nanopore architecture, transiently obstructing ion flow. Importantly, the propensity for gating is not merely dependent on the total charge but intrinsically linked to the exact spatial positioning and polarity of these charges. The research demonstrates that by altering the charge “sign” at specific sites, one can finely tune the nanopore’s gating threshold and conditions, thus redefining the operational stability of these biological conduits.
Complementary experiments also show that reinforcing the structural rigidity of the nanopore abrogates gating behavior entirely. This crucial observation underscores the mechanical flexibility of the pore as a key modulator of gating, shifting the narrative from purely electrostatic considerations to a mechanochemical interplay in ion channel regulation. Such insights open new avenues for engineering nanopores with tailored mechanical properties to either prevent undesirable gating or exploit it for specialized applications.
The implications of these findings extend beyond incremental engineering improvements. The researchers have successfully demonstrated the potential to create nanopores that emulate synaptic plasticity—the brain’s ability to modulate synaptic strength in response to stimuli. By designing nanopores that “learn” from voltage pulses, the team pioneers a bio-inspired computing paradigm that leverages ion flow dynamics for information processing. This revolutionary concept portrays nanopores not just as static sensors but as active components capable of adaptive, memory-like behavior, potentially transforming approaches to neuromorphic computing and ion-based processors.
Exploring the practical applications, this research equips molecular engineers with the knowledge to intentionally circumvent gating in nanopore sensing platforms, thereby enhancing signal stability and measurement accuracy. Conversely, by strategically harnessing gating phenomena, novel classes of ionic devices capable of memory and logic functions can be realized. This dual capability marks a significant leap in the interface between biological nanostructures and advanced computational systems, fostering innovation in biomimetic device architecture.
Supporting institutions involved in this multidisciplinary study include the Institute of Science and Technology Austria, University of Washington, and ENS de Lyon, each contributing expertise critical to experimental design, computational modeling, and theoretical analysis. Their collaboration underscores the global and integrative nature of cutting-edge nanopore research.
The intricacy of ion transport in biological nanopores, long a subject of debate, now rests on a clearer physical foundation thanks to this pioneering work. Through elegant integration of mutation-driven charge reorganization, high-resolution ionic measurements, and theoretical modeling, the diverse and previously mystifying behaviors of nanopores have been coherently demystified. This advancement not only augments the fundamental biophysics of membrane channels but also catalyzes new frontiers in biotechnology, from next-generation DNA sequencers to bio-inspired computing devices.
As nanopore technologies continue their ascent in scientific and technological importance, these insights provide indispensable guidelines for crafting bespoke nanopores with optimized functionalities. The ability to modulate ion transport with such precision embodies a transformative stride toward the full exploitation of biological pores, propelling both our understanding and utilization of nature’s nanoscale machinery.
Subject of Research: Ion transport mechanisms in β-barrel biological nanopores and their biophysical modulation.
Article Title: Lumen charge governs gated ion transport in β-barrel nanopores.
News Publication Date: 11-Nov-2025
Web References:
https://doi.org/10.1038/s41565-025-02052-6
Image Credits: Aleksandra Radenovic/EPFL
Keywords: Biological nanopores, ion transport, ion gating, rectification, aerolysin, β-barrel pore, nanopore sensing, synaptic plasticity mimicry, bio-inspired computing, molecular transport, nanobiophysics.
