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<title>bioRxiv Subject Collection: Biophysics</title>
<link>https://biorxiv.org</link>
<description>
This feed contains articles for bioRxiv Subject Collection "Biophysics"
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<title>bioRxiv</title>
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<link>https://www.biorxiv.org</link>
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<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.29.728470v1?rss=1">
<title>
<![CDATA[
Bayesian Inference of Bond Parameters from Interactions between Single Filaments 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.29.728470v1?rss=1
</link>
<description><![CDATA[
We map interaction forces between two vimentin filaments (cytoskeletal components crucial for cell mechanics) using optical tweezers, while controlling the relative velocity. We introduce a powerful Bayesian inference framework to learn bond parameters directly from force trajectories. The information gained about the bond parameters is maximized by an optimal relative velocity and further by distributing measurements across multiple velocities. Our Bayesian framework is broadly applicable to a large range of biomolecular interactions and force spectroscopy techniques.
]]></description>
<dc:creator><![CDATA[ Pajanonot, K. A. T., Lambert, S., Kumari, P., Koester, S., Klumpp, S. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.29.728470</dc:identifier>
<dc:title><![CDATA[Bayesian Inference of Bond Parameters from Interactions between Single Filaments]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.29.728626v1?rss=1">
<title>
<![CDATA[
Rewiring V-type and K-type enzyme allostery through subunit interface mutations 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.29.728626v1?rss=1
</link>
<description><![CDATA[
Allosteric regulation in the heterodimeric enzyme IGPS depends on long-range communication between the effector-binding HisF subunit and the catalytic HisH subunit. This signaling occurs through a densely packed interdomain interface enriched in conserved noncovalent contacts. Here, we use targeted interface mutations to determine how specific interfacial contacts tune the structural and dynamic features that govern allosteric control. The hK181A variant, which disrupts a critical salt bridge with fD98, converts IGPS into a constitutively more active enzyme, increasing basal glutaminase activity and substrate affinity. By contrast, hR18A, which disrupts a secondary salt bridge with fE71, weakens effector-induced activation, revealing functional asymmetry among interfacial interactions. NMR chemical shift perturbation and CPMG relaxation dispersion experiments show that hK181A remodels millisecond-timescale dynamics throughout HisF, consistent with molecular dynamics simulations indicating enhanced sampling of catalytically competent conformations. Network traffic analysis of correlated communication pathways, combined with energetic analysis, further shows that enthalpic and entropic contributions are redistributed to rewire long-range allosteric signaling. Together, these results identify specific interfacial residues as molecular gates that shape the conformational ensemble accessible to IGPS and show how interface reengineering can be used to rationally reprogram allosteric output.
]]></description>
<dc:creator><![CDATA[ Maschietto, F., Chaudhuri, A., Enny, O., Batista, V. S., Loria, J. P. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.29.728626</dc:identifier>
<dc:title><![CDATA[Rewiring V-type and K-type enzyme allostery through subunit interface mutations]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.29.728618v1?rss=1">
<title>
<![CDATA[
A computational framework to model cartilage degeneration induced by mechanoinflammation and cytokine-driven inflammation in post-traumatic osteoarthritis 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.29.728618v1?rss=1
</link>
<description><![CDATA[
Knee joint injury is a major risk factor for post-traumatic osteoarthritis (PTOA), often associated with early cartilage degeneration. Mechanical overloading and cytokine-driven inflammation are key drivers of this process, yet the underlying mechanisms and their distinct temporal and spatial contributions to cartilage degradation remain unclear. Here, we present a mechanobiological finite element framework that simulates cartilage degradation through cell-mediated proteolytic activity triggered by mechanoinflammation and cytokine-driven inflammation. The model reproduces experimentally observed depth-dependent loss of collagen and aggrecan, with mechanoinflammation inducing a transient response and cytokine-driven inflammation sustaining prolonged matrix degradation. Sensitivity analysis further shows that mechanoinflammation-driven degradation is governed mainly by protease production per cell, whereas cytokine-driven degradation is more sensitive to the rate of cellular stimulation. Together, this framework provides a mechanistic basis to study proteolytic cartilage degeneration and supports future in silico evaluation of therapeutic strategies aimed at mitigating cartilage degradation in PTOA.
]]></description>
<dc:creator><![CDATA[ Hamada, M., Eskelinen, A. S. A., Kosonen, J., Hakonen, S., Florea, C., Grodzinsky, A., Korhonen, R. K., Tanska, P. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.29.728618</dc:identifier>
<dc:title><![CDATA[A computational framework to model cartilage degeneration induced by mechanoinflammation and cytokine-driven inflammation in post-traumatic osteoarthritis]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.06.01.729054v1?rss=1">
<title>
<![CDATA[
Real-time Four-dimensional Imaging and Flow Dynamics Analysis of Cilia-driven Transport in Mammalian Tissues 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.06.01.729054v1?rss=1
</link>
<description><![CDATA[
Quantifying cilia-driven transport in mammalian tissues requires resolving rapid ciliary dynamics while preserving native three-dimensional geometry. Here we combine real-time four-dimensional light-field microscopy with tomographic particle tracking and physics-informed volumetric flow reconstruction to simultaneously image ciliary activity and reconstruct three-dimensional velocity fields in minimally dissected tissues. Applied across brain, respiratory, and reproductive epithelia, the workflow reveals recurrent vortical flow structures and enables quantitative analysis of flow-tissue coupling across individuals.
]]></description>
<dc:creator><![CDATA[ Porcella, G., Sahin, A. T., Keller, J., Nawroth, J. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.06.01.729054</dc:identifier>
<dc:title><![CDATA[Real-time Four-dimensional Imaging and Flow Dynamics Analysis of Cilia-driven Transport in Mammalian Tissues]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.06.01.729299v1?rss=1">
<title>
<![CDATA[
FINDER converts zero-background kinetic fingerprinting into area-scalable attomolar biomarker detection 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.06.01.729299v1?rss=1
</link>
<description><![CDATA[
Background constrains analytical sensitivity: surveying larger sensor areas samples more analyte molecules but also accumulates false positives, limiting gains in detection performance. Here we introduce FINDER[-]Fluorogenic INstantaneous Digital Enumeration and Recognition[-]a single-molecule platform that combines kinetic fingerprinting with fluorogenic transient probes for rapid molecular classification under near-zero-background conditions. By suppressing both solution and surface-associated background at micromolar probe concentrations, FINDER classifies individual molecules within seconds-scale observation windows per field of view. This regime allows sensitivity to scale with surveyed sensor area, enabling amplification-free quantification of the miRNA cancer biomarker hsa-miR-16 with an 11 aM detection limit. FINDER further generalizes to HPV16 DNA biomarker detection, two-color RNA/DNA co-profiling, and rapid discrimination of clinically relevant EGFR single-nucleotide variants using multidimensional kinetic filtering. Rapid per-field classification permits tens of fields to be surveyed within minutes. By converting kinetic specificity into area-scalable sensitivity, FINDER enables semi-automated attomolar biomarker counting without amplification in practical workflows.
]]></description>
<dc:creator><![CDATA[ Walter, N. G., Dai, L., Banerjee, P., Johnson-Buck, A., Blanchard, A., Li, Z. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.06.01.729299</dc:identifier>
<dc:title><![CDATA[FINDER converts zero-background kinetic fingerprinting into area-scalable attomolar biomarker detection]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.31.729144v1?rss=1">
<title>
<![CDATA[
The protein binding domains of staphylococcal protein A fold independently and form an N- to C-terminal gradient of increasing stability. 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.31.729144v1?rss=1
</link>
<description><![CDATA[
Surface factors that contribute to the virulence of Staphylococcus aureus have become therapeutic targets in the treatment of illness associated with this bacterium. Staphylococcal protein A (SpA) is a well-known contributor to S. aureus toxicity and virulence, although relatively little is known about protein A and how its biological function has evolved. SpA is displayed on the surface of the bacterium and contains 5 nearly identical helical ($approx{60}$ aa) domains that bind antibodies with high affinity ($K_dapprox{10}$ nM). The folding free energies of only domains E and B have been determined. In this study we used intrinsic fluorescence detected denaturation to measure the folding thermodynamics of each domain in isolation and in the native multidomain context using a construct that includes the N-terminal half of the mature protein (SpA-N). We also constructed a series of proteins with 1 to 5 repeats of B domain, linked exactly as the five domains of WT SpA are linked. We used nearest neighbor thermodynamic models to explicitly demonstrate that the domains in B domain repeat proteins fold independently. We also showed that the domains in SpA-N fold independently by comparing the folding free energies of domains in isolation and in their multidomain context. Previous dynamic NMR experiments detected highly flexible linkers between domains in 5B, suggesting that the domains of SpA are structurally independent, which is likely responsible for the lack of thermodynamic coupling. Our results also showed a steep increase in domain stability from the N- to C-terminus in SpA-N, from $0.97pm0.05$ to $5.57pm0.28$ kcal/mol. We hypothesize that this stability gradient is related to efficient secretion of protein A.
]]></description>
<dc:creator><![CDATA[ Hagarman, A., Franch, W. R., Oas, T. G. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.31.729144</dc:identifier>
<dc:title><![CDATA[The protein binding domains of staphylococcal protein A fold independently and form an N- to C-terminal gradient of increasing stability.]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.06.01.729284v1?rss=1">
<title>
<![CDATA[
Index-agnostic oblique plane light sheet microscopy of centimetre-scale cleared tissues at subcellular resolution 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.06.01.729284v1?rss=1
</link>
<description><![CDATA[
We present cleared-tissue direct-view oblique plane microscopy (CtDvOPM), which enables optically sectioned subcellular resolution imaging of centimetre-scale tissues at high-throughput over the full range of clearing media refractive indices (n = 1.33-1.56). CtDvOPM can image conventionally-mounted expanded, aqueous or non-aqueous cleared tissue samples at up to 2 m lateral by 14 m axial resolution over a 10 mm x 10 mm x 25 mm sample volume without image tiling, at up to 400 million voxels per second.
]]></description>
<dc:creator><![CDATA[ Lamb, J. R., Cardoso Mestre, M., Fenwyn Longrin, K., Bhat, P., Stevenson, M., Rhodes, A. D. Y., Gosieniecka, J., Redmond, L. C., Higgins, C. A., Rodriguez-Rodrigues, N., Lancaster, M. A., Manton, J. D. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.06.01.729284</dc:identifier>
<dc:title><![CDATA[Index-agnostic oblique plane light sheet microscopy of centimetre-scale cleared tissues at subcellular resolution]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.06.01.729289v1?rss=1">
<title>
<![CDATA[
A self-consistent model for phase separation and active processes in biomolecular condensates 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.06.01.729289v1?rss=1
</link>
<description><![CDATA[
Biomolecular condensates are thought to play a pivotal role in cellular organization by regulating biochemical reactants in space and time. Sustained molecular fluxes across condensate boundaries, together with the participation of phase-separating molecules in active chemical reactions such as ATP hydrolysis, call for a nonequilibrium description. Here, we propose a self-consistent framework in which diffusion--drift dynamics and chemical reactions are coupled through a conditional free energy, defined as the excess contribution to the chemical potential. Self-consistency is achieved by deriving this quantity from the same free-energy functional that governs molecular interactions and phase separation. We apply the framework to a minimal client--scaffold system and investigate how active chemical processes and phase separation interact at steady state. In doing so, our approach recovers the fundamental rules previously identified for the emergence of nonequilibrium steady-state fluxes. The model shows that active reactions involving the scaffold molecules can regulate the phase behavior of the condensate. Moreover, nonequilibrium steady-state fluxes are maximal near the boundary between the phase-separated and homogeneous regimes, suggesting that condensates sustaining molecular transport may operate close to their stability threshold. In the same region, client fluxes are also enhanced, revealing an indirect coupling between scaffold activity and client transport. These results provide a baseline for developing more detailed theories of chemically active condensates.
]]></description>
<dc:creator><![CDATA[ Di Mambro, M., De Los Rios, P. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.06.01.729289</dc:identifier>
<dc:title><![CDATA[A self-consistent model for phase separation and active processes in biomolecular condensates]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.31.729125v1?rss=1">
<title>
<![CDATA[
Fingertip real contact area scales quadratically with input voltage in electrostatic actuation 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.31.729125v1?rss=1
</link>
<description><![CDATA[
Touchscreens have become the dominant interface in consumer electronics, yet interactions with them remain primarily visual. Incorporating haptic feedback that simulates touch sensations could make these interactions more natural and intuitive. Electrostatic actuation, which modulates friction by attracting the finger toward a capacitive surface using an alternating voltage, offers a promising approach. The resulting increase in friction is often attributed to the rise in real contact area; however, direct experimental evidence linking voltage input parameters to real contact area and contact forces remains limited. Here, we use frustrated total internal reflection to directly image the real contact area while simultaneously measuring contact forces during controlled finger sliding under electrostatic actuation. We systematically vary voltage amplitude (75-150 V) and excitation frequency (30-230 Hz) and quantify the changes in contact area and forces as functions of these parameters. Our results reveal a quadratic dependence of real contact area, electrostatic attraction, and tangential force on voltage amplitude, with comparatively small effects of excitation frequency. These findings clarify the respective roles of voltage amplitude and frequency in the electrostatic modulation of finger contact mechanics, providing design guidelines for haptic display design.
]]></description>
<dc:creator><![CDATA[ Kenanoglu, C. U., Vardar, Y. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.31.729125</dc:identifier>
<dc:title><![CDATA[Fingertip real contact area scales quadratically with input voltage in electrostatic actuation]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.06.01.729173v1?rss=1">
<title>
<![CDATA[
Hashi: Bridging Statistical Model Derived 1D Microstate Encodings and Protein 3D Structural Ensembles 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.06.01.729173v1?rss=1
</link>
<description><![CDATA[
The functioning of proteins is intimately linked to the conformational states they sample within the native ensemble. Generating ensembles from a single static structure is therefore a research domain receiving considerable attention. In this application note, we introduce Hashi, a pipeline to rapidly generate realistic structural ensembles from the outputs of the structure-based Wako-Saito-Munoz Eaton (WSME) statistical mechanical model of protein folding. This approach relies on integrating the block WSME model outputs - strings of zeros and ones describing the conformational status of every residue over thousands or millions of microstates each assigned a statistical weight derived from physically grounded energy-entropy terms, and free energy profiles - with the RANCH module of the EOM (ensemble optimization method) from the ATSAS software suite, providing three-dimensional views of the structural ensembles within the model framework. It is applicable to a variety of single-chain monomeric systems with lengths ranging from 30 to 500 residues, including globular and repeat proteins. The generated structural ensembles can also be rank ordered according to their free energies within a given macrostate or a range of reaction coordinate values. Since the statistical weights of the WSME model microstates can be reweighted or calibrated with experiments, the ensembles shed light on not just the folding mechanism but also on the structural excursions that determine function and opening of otherwise buried binding pockets.
]]></description>
<dc:creator><![CDATA[ Naganathan, A. N., Madhan, H. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.06.01.729173</dc:identifier>
<dc:title><![CDATA[Hashi: Bridging Statistical Model Derived 1D Microstate Encodings and Protein 3D Structural Ensembles]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.29.728696v1?rss=1">
<title>
<![CDATA[
Physics-guided design of intrinsically disordered proteins 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.29.728696v1?rss=1
</link>
<description><![CDATA[
Intrinsically disordered protein regions (IDPs) are found across the tree of life and characterized by the lack of a stable 3D fold, encoding function through a vast ensemble of conformations. This plasticity makes rational design of IDPs challenging. Physics-based approaches capturing distinct aspects of sequence composition, charge patterning, and molecular interactions have emerged as powerful predictors of ensemble-derived properties. Here, we present a machine learning framework for proteome-scale de novo IDP design by rationally inverting physics-based models. We first program IDPs to tunably sense and respond to diverse biophysical cues and show that IDP ensembles can directly encode complex signal processing, including threshold detection, bandpass filtering, and Boolean-type multi-input logic. We next engineer multicomponent IDP mixtures with tailored emergent condensate properties, including layering and number of phases, compositional specificity, and RNA-dependent remodeling of structure and composition. Finally, we demonstrate designed IDPs that selectively partition into or deplete from biological condensates in living cells. Together, our framework establishes a flexible and scalable strategy for design of ensemble-derived and collective properties in dynamic biomolecules.
]]></description>
<dc:creator><![CDATA[ Tyagi, N., Boodry, J., Chou, V., Snead, W. T., Shrinivas, K. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.29.728696</dc:identifier>
<dc:title><![CDATA[Physics-guided design of intrinsically disordered proteins]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.29.728729v1?rss=1">
<title>
<![CDATA[
Synthetic Fibrous Hydrogels as Minimal Systems to Modulate Cell Migration Modes in 3D 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.29.728729v1?rss=1
</link>
<description><![CDATA[
Cell migration in three-dimensional (3D) environments is highly plastic and regulated by extracellular matrix (ECM) cues. Engineered biomaterials provide controllable platforms to investigate how specific matrix signals regulate cell behavior in 3D, yet how defined biochemical signals control migration modes remain unclear. Here, we present tunable fibrous polyisocyanide (PIC) hydrogels functionalized with integrin-binding RGD peptides, cadherin-mimetic HAVDI peptides, or no ligands to direct mesenchymal, hybrid, or amoeboid-like migration of human adipose-derived stem cells without altering matrix mechanics. Using live-cell tracking, 3D displacement microscopy, matrix remodeling analysis, and YAP nuclear localization, we show that ligand identity governs adhesion organization, force transmission, and mechanotransduction. RGD-functionalized matrices promote {beta}1-integrin clustering, extensive matrix remodeling, strong YAP activation and upregulation of migration-related genes. In contrast, non-adhesive matrices limit adhesion formation, resulting in weak force transmission and amoeboid-like behavior. HAVDI-functionalized matrices induce cadherin clustering and heterogeneous cellular responses, indicating that a hybrid migration mode arises from adhesion organization rather than a distinct transcriptional program. Together, these findings demonstrate that ligand identity alone is sufficient to program migration mode in a force-responsive 3D matrix and provide a versatile platform to dissect cell-matrix interactions in complex environments.
]]></description>
<dc:creator><![CDATA[ Zhang, H., Solis Fernandez, G., Louis, B., Vorsselmans, S., Hofkens, J., Kouwer, P. H. J., Yuan, H., Rocha, S. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.29.728729</dc:identifier>
<dc:title><![CDATA[Synthetic Fibrous Hydrogels as Minimal Systems to Modulate Cell Migration Modes in 3D]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.30.729014v1?rss=1">
<title>
<![CDATA[
Extracting anomalous diffusion parameters from multi-state ensembles of short single molecule trajectories. 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.30.729014v1?rss=1
</link>
<description><![CDATA[
Single-molecule tracking measures the stochastic motion of individual biomolecules in the cellular environment. Statistical analysis of trajectory ensembles is required to gain insight into the biophysical nature of mobility states and molecular interactions that they reflect. Mobility states can be parameterized by a generalized diffusion coefficient and anomalous exponent. Experimental constraints such as finite track length and localization precision limit how accurately these parameters can be determined. We compare the performance of analysis methods to recover the input parameters from ensembles of simulated single molecule tracks from different states spanning the range of anomalous diffusive behaviors observed in the cell nucleus. We further develop a framework to quantify error rates in the assignment of mobility states to individual molecules based on recall rates and precision. Our analysis shows that single-track analysis methods are superior to bulk methods in their ability to recover parametric descriptors from mixed populations. The most complete description is obtained by combining outputs from different tools. Our work provides a guide to assess the accuracy of analyses and obtain the most accurate parametric description of experimental single particle tracking data.
]]></description>
<dc:creator><![CDATA[ Budhathoki, A., Pandey, G., Galeota-Sprung, J., Spille, J.-H. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.30.729014</dc:identifier>
<dc:title><![CDATA[Extracting anomalous diffusion parameters from multi-state ensembles of short single molecule trajectories.]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.29.728850v1?rss=1">
<title>
<![CDATA[
Spin-Dependent Extracellular Respiration 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.29.728850v1?rss=1
</link>
<description><![CDATA[
Biological energy conversion relies on highly efficient electron transfer. The chirality induced spin selectivity (CISS) effect, which couples electron spin to momentum in chiral molecules, is hypothesized to promote this efficiency. While observed in isolated biomolecules, the physiological relevance of CISS during active cellular metabolism remains unknown. Here, we demonstrate that CISS influences extracellular electron transfer in living Geobacter sulfurreducens biofilms. Cultivation on ferromagnetic electrodes yields a significant asymmetry in respiratory current between opposite substrate spin states. Furthermore, in situ magnetization reversal induces reversible changes in respiratory flux. These results provide the first in vivo demonstration that spin selectivity directly impacts respiration. By revealing a quantum feature of extracellular respiration, our findings offer a strategy to exploit the spin degree of freedom in bioelectronics.
]]></description>
<dc:creator><![CDATA[ Sukenik, N., Harris, C. C., Yadav, S., Chavez, M. S., Niman, C. M., Baczewski, L. T., El-Naggar, M. Y. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.29.728850</dc:identifier>
<dc:title><![CDATA[Spin-Dependent Extracellular Respiration]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.31.729094v1?rss=1">
<title>
<![CDATA[
Single-Cell Electrophysiology Reveals Verapamil's Disruption of Bacterial Membrane Energetics 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.31.729094v1?rss=1
</link>
<description><![CDATA[
Verapamil, a clinically used calcium channel blocker, enhances the activity of several tuberculosis antibiotics, but its mechanism of action and physiological effects on bacteria remain unresolved. A central debate concerns whether verapamil primarily inhibits efflux pumps or disrupts membrane energetics. Here, we use Escherichia coli as a model system to quantify single-cell and population-level physiological responses to verapamil with high temporal resolution. Real-time measurements of the rotational speed of individual flagellar motors, a single-cell proxy for the proton motive force (PMF), reveal a heterogeneous response to verapamil: treated cells exhibit either a dose-dependent gradual decrease in PMF, or a rapid collapse of PMF. Although loss of the outer-membrane efflux channel TolC increases growth inhibition by verapamil, it does not alter the rapid PMF disruptions observed at the single-cell level, suggesting that efflux contributes to long-term susceptibility but not to the initial PMF disruption. Independent assays of population-level motility, pH, and membrane-integrity suggest that verapamil may selectively dissipate the electrical component of PMF while leaving intracellular pH largely unchanged. A minimal electrical circuit model captures both steady-state and dynamic behavior. Together, these findings demonstrate that verapamil rapidly and reversibly perturbs bacterial membrane energetics through a mechanism distinct from classical protonophores, helping to reconcile conflicting interpretations of its activity and clarifying how membrane effects may interact with efflux inhibition during antibiotic potentiation.
]]></description>
<dc:creator><![CDATA[ Biquet-Bisquert, A., Astezan, A., Marmol, M., Voyvodic, P. L., Mohite, N., Pedaci, F., Nord, A. L. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.31.729094</dc:identifier>
<dc:title><![CDATA[Single-Cell Electrophysiology Reveals Verapamil's Disruption of Bacterial Membrane Energetics]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.29.728845v1?rss=1">
<title>
<![CDATA[
Multimodal dynamics control activity of a glial glutamate transporter 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.29.728845v1?rss=1
</link>
<description><![CDATA[
Membrane transporters move polar solutes across lipid bilayers to regulate cellular metabolism, signaling, and drug distribution. These proteins operate via an alternating-access mechanism, cycling between extracellular-, intermediate-, and intracellular-facing conformations. The human excitatory amino acid transporter 1 (EAAT1) protects neurons from excitotoxic damage by mediating the uptake of glutamate and aspartate into glial cells. Defects in EAAT1 function result in numerous pathologies, including epilepsy and ataxia, suggesting that positive modulation of these transporters might ameliorate glutamate neurotoxicity. However, developing EAAT1 activators requires understanding the timing of conformational changes, which remain largely unexplored. Here, we establish an experimental platform that combines single-molecule Forster resonance energy transfer (smFRET) to monitor real-time conformational dynamics, single-transporter activity assays to correlate dynamics with function, and cryogenic electron microscopy (cryoEM) to visualize discrete conformations at high resolution. This platform enables detection of Angstrom-scale movements of single transporter molecules in real time, revealing that EAAT1 intersperses rapid conformational dynamics with long pauses. Slow and fast dynamics can be modulated by substrates, membrane composition, and mutations, and are correlated with the enrichment of specific structural states. We leverage this platform to investigate an EAAT1 mutation associated with severe episodic ataxia and show that it inhibits transport by stabilizing a paused cytoplasm-facing conformation. These results identify multimodal dynamics as an intrinsic, regulatable feature of EAAT1 function and, therefore, a potential therapeutic target. Henceforth, our integrated platform will facilitate investigations of other regulatory factors, including the effects of small-molecule and lipid modulators on the transport cycle.
]]></description>
<dc:creator><![CDATA[ Wu, Q., Ciftci, D., Canul Tec, J., Reyes, N., Gregorio, G., Huang, Y., Boudker, O. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.05.29.728845</dc:identifier>
<dc:title><![CDATA[Multimodal dynamics control activity of a glial glutamate transporter]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.06.01.729260v1?rss=1">
<title>
<![CDATA[
Single-molecule visualisation of human Hsp70-driven conformational remodelling during stress 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.06.01.729260v1?rss=1
</link>
<description><![CDATA[
The Hsp70 chaperone system plays a central role in the folding of nascent polypeptides and in preventing protein misfolding and aggregation during cellular stress. However, the precise mechanism by which the human Hsp70, HspA8, remodels the conformations of individual chemically misfolded clients remains unclear. Moreover, it is unknown whether this mechanism differs from that used by Hsp70 to engage clients during heat stress to preserve native function. To address these questions, we report here the use of single-molecule fluorescence resonance energy transfer (smFRET) to temporally interrogate how the human Hsp70 system regulates the conformation of a heat-sensitive client protein, firefly luciferase (Fluc), following chemical denaturation and during heat stress. We find that Hsp70 recognises both chemically denatured and heat-induced misfolded states of Fluc and resolves them by conformational expansion. Release from a Hsp70-bound state, a process driven by the nucleotide exchange factor, Hsp110, guides Fluc toward productive folding trajectories that would otherwise be unlikely to occur spontaneously following collapse from a conformationally unfolded state. Moreover, we demonstrate that both temperature and the conformational state of misfolded Fluc dictate the ability of HspA8 to meaningfully resolve non-native structure within the protein. Collectively, this work provides direct visualisation of the mechanisms by which Hsp70 modulates client conformations under diverse stress conditions to preserve proteome integrity.
]]></description>
<dc:creator><![CDATA[ Skewes, B., McMahon, S., Auld, N. K., Marzano, N. R., van Oijen, A. M., Ecroyd, H. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.06.01.729260</dc:identifier>
<dc:title><![CDATA[Single-molecule visualisation of human Hsp70-driven conformational remodelling during stress]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.06.01.729272v1?rss=1">
<title>
<![CDATA[
Simulation-driven discovery of morphology-function relationships in microswimmers 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.06.01.729272v1?rss=1
</link>
<description><![CDATA[
For more than a billion years, microorganisms have evolved complex strategies for navigating aquatic habitats, despite the fundamental limitations and constraints imposed by their physical environment. A common theme across these strategies is the use of active slender appendages (cilia, flagella, archaella) to generate self-propulsion. Diverse selection pressures and evolutionary trajectories have driven the emergence of drastically different morphologies of biological microswimmers, each tailored for distinct functions ranging from motility to taxis to prey capture to feeding. Despite the biological and ecological significance of these intricate microscale processes, realistic computational modelling of these organisms and their behaviours is still in its infancy. Here, we present a comprehensive open-source simulation platform for motile microswimmers, that faithfully captures the universal hydrodynamic principles shared by such systems. We illustrate the predictive power and versatility of this approach to resolve and explore morphology-function relationships across different microswimmer species and provide new insights into the diversification of locomotion strategies in early eukaryotes.
]]></description>
<dc:creator><![CDATA[ Cass, J. F., Wan, K. Y. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.06.01.729272</dc:identifier>
<dc:title><![CDATA[Simulation-driven discovery of morphology-function relationships in microswimmers]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.06.01.729280v1?rss=1">
<title>
<![CDATA[
Quantitative biophysical analysis of human septin hexamer and octamer self-assembly on model membranes 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.06.01.729280v1?rss=1
</link>
<description><![CDATA[
Septins are GTP-binding cytoskeletal proteins that shape and compartmentalize the plasma membrane. Their complex interactome has made it difficult to understand the molecular factors that govern their assembly. Moreover, it is unclear whether human septin hexamers and octamers form distinct higher-order assemblies, especially at the plasma membrane. Here, we address this question by using label-free methods to probe binding and self-assembly of purified human septins on supported lipid bilayers. Quartz crystal microbalance with dissipation (QCM-D) monitoring revealed that septin-membrane binding is diffusion-limited and concentration-dependent. The viscoelastic properties differed between septin oligomers suggestive of structural differences. Imaging of membrane-bound septin networks by atomic force microscopy (AFM) revealed that septin hexamers formed aligned nematic filamentous networks, whereas septin octamers formed aligned curved structures including spirals. QCM-D and AFM measurements both showed that septins form double-layered filament networks. However, upon C-terminal truncation of the SEPT6 and SEPT7 subunits, hexamers no longer bound the membrane while octamers formed a single-layered network of filament spirals. Our findings reveal that human septin hexamers and octamers interact differently with membranes, providing a baseline to understand their functions in the cell.
]]></description>
<dc:creator><![CDATA[ Reese, S., de Ridder, W., van Hemmen, A., Mateescu, A.-G., Togo, R., Omi, S., Mavrakis, M., Richter, R., Koenderink, G. H. ]]></dc:creator>
<dc:date>2026-06-02</dc:date>
<dc:identifier>doi:10.64898/2026.06.01.729280</dc:identifier>
<dc:title><![CDATA[Quantitative biophysical analysis of human septin hexamer and octamer self-assembly on model membranes]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-02</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.30.729023v1?rss=1">
<title>
<![CDATA[
Do AI Structure Predictors Capture Bound-State Disorder? A Benchmark on Fuzzy Protein Complexes 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.30.729023v1?rss=1
</link>
<description><![CDATA[
Fuzzy protein complexes, in which an intrinsically disordered protein (IDP) retains conformational disorder upon binding, pose a fundamental challenge for structure predictors trained on ordered systems, where crystal structures capture only the most ordered ensemble snapshot, making standard benchmarking metrics misleading. Here, we present the first systematic evaluation of AlphaFold3 (AF3), AlphaFold2-Multimer (AF2MM), Chai-1, and Boltz-2 on a curated dataset of fuzzy complexes from FuzDB, benchmarked against DockQ against PDB structures and NOE violation rates against manually curated BMRB restraint files, the first comprehensive collection of this kind. Across all four predictors, approximately 30% of NOE restraints were violated with nearly identical distributions regardless of predictor architecture or training data. DockQ scores fell uniformly in the Acceptable range, with AF3 marginally higher but showing equivalent NOE violation rates to the weakest-performing model. Ensemble-level analysis using a first-principles implementation of the Hadzi thermodynamic model revealed that AF3 uniquely achieves near-zero mean helicity bias versus systematic overconfidence in the other predictors, yet all four models show poor per-residue helicity correlation with thermodynamic expectations. DockQ rankings reflect training data similarity to crystal structures rather than physical accuracy, and no current predictor captures fuzzy complex ensemble behavior. The FuzzyBench-NOE dataset, comprising NOE restraint files, predicted structures, interface hotspot annotations, and Hadzi--DSSP analysis outputs, is released on Zenodo (https://doi.org/10.5281/zenodo.20470556).
]]></description>
<dc:creator><![CDATA[ Velasquez, J., Ghent, S., Rahman, T. ]]></dc:creator>
<dc:date>2026-06-01</dc:date>
<dc:identifier>doi:10.64898/2026.05.30.729023</dc:identifier>
<dc:title><![CDATA[Do AI Structure Predictors Capture Bound-State Disorder? A Benchmark on Fuzzy Protein Complexes]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.28.728436v1?rss=1">
<title>
<![CDATA[
Local mechanical gradients underlie coordinated cascades of epithelial cell-cycle advance 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.28.728436v1?rss=1
</link>
<description><![CDATA[
Cell division mechanically perturbs the local environment in epithelial tissues, yet whether and how these perturbations propagate to coordinate cell-cycle progression across neighboring cells remains unclear. Here, we combine live cell-cycle tracking with mechanical analysis to examine how division events organize in space and time. We find that coordinated changes and spatial gradients in forces, morphology, and dynamics precede synchronized G1[-&gt;]S transitions in nearby cells. These transitions emerge within localized high-tension zones and give rise to spatiotemporal clusters of cell divisions, indicating that division events are mechanically coupled and propagate across neighboring cells. Supporting this biophysical picture, similar mechanical patterns arising from cell extrusion are sufficient to induce cell-cycle re-entry in neighboring cells. Together, these findings suggest a mechanically mediated framework for coordinated proliferation, possibly driven by positive mechanical feedback.
]]></description>
<dc:creator><![CDATA[ Wanszelbaum, S., Saleem, A., Daraf, L., Lavi, Y., Atia, L. ]]></dc:creator>
<dc:date>2026-06-01</dc:date>
<dc:identifier>doi:10.64898/2026.05.28.728436</dc:identifier>
<dc:title><![CDATA[Local mechanical gradients underlie coordinated cascades of epithelial cell-cycle advance]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.27.727973v1?rss=1">
<title>
<![CDATA[
Electrokinetic active grooves for liposome capture, confinement, trajectory analysis and controlled release 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.27.727973v1?rss=1
</link>
<description><![CDATA[
Extracellular vesicles (EVs) are attractive candidates for minimally invasive disease monitoring. EV diagnostic signals often reside in low-abundant subpopulations, motivating single-vesicle measurements that can report distributions rather than ensemble averages. In practice, such assays rely on transient migration of a mobile vesicle through a detection region or performing cycled processing/measurement steps on vesicles permanently immobilized on surfaces via affinity-based capture. Single vesicle analysis would benefit from technologies that can combine characteristics of transient detection and surface immobilization to enable controlled capture, fixed time analysis and release of vesicles in a marker-neutral way. Here, we present electrokinetically active (RECON) grooves that provides low-voltage, programmable capture, timed retention, and release of nanoscale vesicles within microscopic grooves under continuous optical access. The platform uses a titanium embedded electrode with ultrathin TiO2 passivation, enabling stable actuation over multi-hour experiments. Using fluorescent, size-defined liposomes as standards, we demonstrate (1) reversible, minute-long confinement at <5 V, (2) size-dependent confinement times, and (3) optical quantification of groove-guided drift velocities. Together, these results establish RECON grooves as a reusable, electrically programmable platform for controlled capture, timed retention, and release, while providing quantitative, trajectory-derived measures of confinement and transport.
]]></description>
<dc:creator><![CDATA[ Ravikumar, P., Pessoa, M. A. S., Shiekh, S., Liu, Z., Sladek, R., Reisner, W. ]]></dc:creator>
<dc:date>2026-06-01</dc:date>
<dc:identifier>doi:10.64898/2026.05.27.727973</dc:identifier>
<dc:title><![CDATA[Electrokinetic active grooves for liposome capture, confinement, trajectory analysis and controlled release]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.28.728368v1?rss=1">
<title>
<![CDATA[
MEMBRANE PORES ACT AS SELF-RESEALING LIPID SCRAMBLASES 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.28.728368v1?rss=1
</link>
<description><![CDATA[
Biological membranes continuously experience leaflet lipid imbalances during growth, lipid synthesis, and vesicle fusion. To alleviate such imbalances and prevent these asymmetries from compromising membrane integrity, cells rely on lipid scramblases, fast and non-specific lipid channel proteins. Here we show that lipid number asymmetry alone is sufficient to drive spontaneous formation of transient hydrophilic pores that function as self-resealing lipid scramblases. Using giant unilamellar vesicles, living cells, and coarse-grained molecular dynamics simulations, we demonstrate that fusion-induced excess lipids in one leaflet lowers membrane edge tension, generating size-selective pores whose size and lifetime scale with the magnitude of asymmetry. Below a critical threshold, these pores reseal spontaneously; above it, membranes collapse. Strikingly, pore opening enables rapid, non-selective lipid translocation between leaflets, dissipating the asymmetry that nucleates the pore and thereby promoting their own closure. Cholesterol buffers moderate imbalances through spontaneous flip flop before pore formation, whereas pore-mediated lipid scrambling relieves the remaining asymmetry and restores cholesterol's initial distribution. Our findings identify transient lipid pores as an intrinsic, protein-independent mechanism that couples membrane destabilization to self-repair, providing a universal physical principle for membrane homeostasis during growth, remodelling, and early cellular evolution.
]]></description>
<dc:creator><![CDATA[ Lira, R. B., van Tilburg, M., Cavalcanti, R., Richards, C., Riske, K., Marrink, S.-J., Roos, W. H. ]]></dc:creator>
<dc:date>2026-06-01</dc:date>
<dc:identifier>doi:10.64898/2026.05.28.728368</dc:identifier>
<dc:title><![CDATA[MEMBRANE PORES ACT AS SELF-RESEALING LIPID SCRAMBLASES]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.28.728474v1?rss=1">
<title>
<![CDATA[
Network-Level Characterization of Spontaneous Calcium Activity in an In-Vitro Alzheimer's Disease Model 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.28.728474v1?rss=1
</link>
<description><![CDATA[
The neurodegenerative disorder Alzheimer's disease (AD) is widely known for biomarkers such as amyloid beta plaques and tauopathy, as well as functional differences in memory and cognitive ability. Despite this devastating functional impact, a large body of work only focuses on molecular biomarkers of AD. In this study, we investigate collective neural dynamics in vitro and assess how network-level properties differ between a well-established model of familial AD (FAD) and a newly developed in vitro accelerated model (acAD). The new model system reliably develops the key structural characteristics of AD in three weeks, but its calcium dynamics had not been characterized previously. Spontaneous network dynamics influences information processing as part of the internal network state. Here we measure this spontaneous activity of a network of hundreds of cells in each field of view. We find that the FAD model has a larger fraction of hyperactive cells, while the acAD model displays similar characteristics to healthy cells. Additionally, the FAD model has altered cooperation between cells, losing a proportion of highly correlated cellular activities, both for fast and slow coupling among cells. The acAD model is again consistent with healthy networks. Since the acAD model does not show the same spontaneous network dysfunction seen in FAD, it can enable measurements of changes in learning and memory associated with the plasticity, rather than the structure of the internal network state.
]]></description>
<dc:creator><![CDATA[ Emenheiser, A. M., Gentry, E., Xue, H., Alvarez, P., O'Neill, K., Cao, K., Losert, W. ]]></dc:creator>
<dc:date>2026-06-01</dc:date>
<dc:identifier>doi:10.64898/2026.05.28.728474</dc:identifier>
<dc:title><![CDATA[Network-Level Characterization of Spontaneous Calcium Activity in an In-Vitro Alzheimer's Disease Model]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.28.728353v1?rss=1">
<title>
<![CDATA[
A Charge Detection Mass Spectrometer for the Analysis of Megadalton-sized Molecules 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.28.728353v1?rss=1
</link>
<description><![CDATA[
Advances in Electrostatic Linear Ion Trap (ELIT) Charge Detection Mass Spectrometry (CDMS) over the past 10 years have revolutionized its use for analyzing very high-molecular-weight species such as protein complexes, viral vectors, vaccines, viruses, and amyloid fibrils. Nonetheless, ELIT-based CDMS has remained confined to a small number of specialized instrumentation groups, predominantly in academia, where large and complex home-built instruments are operated by highly skilled scientists in dedicated facilities. In this report, we discuss the primary challenges addressed in the design of a benchtop ELIT-based CDMS instrument. We highlight key design aspects of the hardware, acquisition modes, and control software, and we present important performance metrics (mass range, resolution and sensitivity) demonstrated using samples representative of the technology's key application areas.
]]></description>
<dc:creator><![CDATA[ Ujma, J., Wheeldon, C., Schofield, A., Danby, M., Eatough, D., Bruton, D., Haris, A., Richardson, K., Langridge, D., Jarrell, A., Brown, J. M., Draper, B. E., Jarrold, M. F., Giles, K. ]]></dc:creator>
<dc:date>2026-06-01</dc:date>
<dc:identifier>doi:10.64898/2026.05.28.728353</dc:identifier>
<dc:title><![CDATA[A Charge Detection Mass Spectrometer for the Analysis of Megadalton-sized Molecules]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.28.728365v1?rss=1">
<title>
<![CDATA[
A Closed-Form Bayesian Framework for DNA Replication Reveals Intrinsic Origin Timing and Activation Delays 
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</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.28.728365v1?rss=1
</link>
<description><![CDATA[
We present an analytical framework for modeling eukaryotic DNA replication that, given experimental Replication Fork Directionality (RFD) data, enables Bayesian inference of origin number, activation delay (t) and intrinsic timing ({lambda}), the mean replication time if each origin were isolated. By deriving closed-form expressions for RFD and Mean Replication Timing (MRT) under exponential and a specific Weibull firing-time distributions as functions of (t) and ({lambda}), we eliminate the need for stochastic simulations. These analytical results reveal that RFD, as a ratio of fork directions, is invariant under joint rescaling of intrinsic timing and fork speed; absolute intrinsic timing can nonetheless be inferred when fork speed is independently measured. We demonstrate that under exponential firing distribution for the origin, the observed efficiency (E), i.e. the probability for an origin to fire which accounts for nearby origins, is simply MRT(x)/{lambda}. The closed-form RFD expressions allow use of a Bayesian method that achieves 0.96-0.99 correlation with yeast RFD profiles and resolves [~]780 origins in S. cerevisiae. Our framework identifies about 150 origins with biologically significant delays ([&ge;] 3 minutes), revealing regulated activation kinetics undetectable by existing methods. By quantifying how origin intrinsic timing and delays shape replication timing landscapes, this work confirms yeast as a paradigm organism for studying DNA replication control mechanisms.
]]></description>
<dc:creator><![CDATA[ D'Asaro, D., Ciardo, D., Hyrien, O., Lacroix, L., Le tallec, B., Goldar, A., Audit, B., Arbona, J.-M. ]]></dc:creator>
<dc:date>2026-06-01</dc:date>
<dc:identifier>doi:10.64898/2026.05.28.728365</dc:identifier>
<dc:title><![CDATA[A Closed-Form Bayesian Framework for DNA Replication Reveals Intrinsic Origin Timing and Activation Delays]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.28.728492v1?rss=1">
<title>
<![CDATA[
Lipids intercalate and mediate multi-channel assemblies of connexin-46/50 gap junctions 
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</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.28.728492v1?rss=1
</link>
<description><![CDATA[
Gap junction channels enable direct electrical and metabolic exchange between adjacent cells and tissues, where they organize into dense plaques containing tens to thousands of channels. Although plaque formation is known to modulate junctional conductance, the structural basis for channel-channel organization within a membrane environment remains poorly defined. Here, we reconstitute native lens connexin-46/50 (Cx46/50) gap junction channels into MSP-based lipid nanodiscs that incorporate multiple channels to create miniature plaque-like complexes suitable for single-particle cryo-electron microscopy (cryo-EM). We determine high-resolution structures of dual-channel assemblies in two distinct configurations and find no ordered protein-protein contacts across the interface. Instead, the inter-channel space is occupied by ordered lipid density, indicating that channel packing in these assemblies is lipid mediated. These channel-channel interfaces stabilize discrete lipid populations, including an interstitial lipid intercalated between subunits and positioned near the N-terminal gating domain, suggesting a route by which channel organization could promote lipid occupancy near the pore. Additionally, we leverage this dataset to refine the Cx46/50 single-channel structure to 1.8 [A] resolution, revealing exceptional chemical detail of the pore-lining landscape in the stabilized open-state. Together, these results define principles of lipid-mediated multi-channel organization and suggest how plaque-like packing may tune gap junction function through specific lipid interactions.
]]></description>
<dc:creator><![CDATA[ Garrels, C. S., Myers, J. B., Souza, S. A., Jarodsky, J. M., Reichow, S. L. ]]></dc:creator>
<dc:date>2026-06-01</dc:date>
<dc:identifier>doi:10.64898/2026.05.28.728492</dc:identifier>
<dc:title><![CDATA[Lipids intercalate and mediate multi-channel assemblies of connexin-46/50 gap junctions]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-06-01</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.27.728205v1?rss=1">
<title>
<![CDATA[
Antibody maturation increases rigidity in protein-contacting regions and flexibility at glycan interfaces 
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</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.27.728205v1?rss=1
</link>
<description><![CDATA[
Antibody design is a challenging task that could be improved by understanding the conformational changes accompanying affinity maturation. Antibody maturation is a critical immune system process by which antibodies gain mutations that improve affinity and specificity for antigen targets. However, how these mutations change antibody dynamics, specifically whether antibodies become more rigid as they mature, remains a contested topic. Using adaptive sampling molecular dynamics simulations and over 8.5 milliseconds of Folding@home simulations of seven lineages, we discover that affinity maturation selectively tunes the dynamics of antibody paratope regions depending on whether they contact protein residues or glycans. We find that antibody regions that contact glycans evolve to become more flexible and, consistent with other studies, antibody regions that contact protein residues on the antigen become rigid. This pattern holds regardless of whether one includes the constant region in the simulations, indicating that the computational cost of all-atom antibody MD simulations can be reduced by half without sacrificing the accuracy of the variable region dynamics. We expect the principles identified in this study will enable precise, dynamics-based engineering of high-affinity antibodies and to inform immunogen design against challenging, glycan-shielded viral targets such as HIV.
]]></description>
<dc:creator><![CDATA[ Solieva, S. O., Park, J., Miller, J. J., Kriews, A., Escolano, A., Bowman, G. R. ]]></dc:creator>
<dc:date>2026-05-31</dc:date>
<dc:identifier>doi:10.64898/2026.05.27.728205</dc:identifier>
<dc:title><![CDATA[Antibody maturation increases rigidity in protein-contacting regions and flexibility at glycan interfaces]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-31</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.29.728716v1?rss=1">
<title>
<![CDATA[
Electrically programmable picoscale phototransduction of a newly discovered microbial rhodopsin 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.29.728716v1?rss=1
</link>
<description><![CDATA[
Human retina can achieve single-photon sensitivity through specialised photoreceptors that convert light into electrical signals via phototransduction. Among microbial light-sensitive proteins, proteorhodopsins stand out for their intrinsic light-driven ion transport and spectral tunability, making them promising candidates for bio-inspired photonic devices. A central challenge for acellular integration, however, is the fragility of most bacterial rhodopsins under extreme conditions. Here, we exploit the exceptional robustness of TARA76, a microbial rhodopsin that retains structural integrity even upon complete dehydration, to demonstrate its functional reconstitution in an artificial black lipid membrane within a biocompatible microfluidic platform. By recording light-induced ionic currents with picoampere sensitivity across a broad range of pH, illumination power, electrolyte composition, and applied voltages, we establish TARA76 as a high-performance photoelectric transducer in a fully acellular environment. Strikingly, we uncover a strong and previously unreported dependence of the photocurrent on Na ions, which appears to play a key structural and functional role in stabilising the protein's active conformation. Furthermore, we demonstrate that the orientation of TARA76 within the artificial membrane can be externally controlled by applying a defined electric field during bilayer formation, enabling deterministic tuning of photocurrent directionality. Together, these results establish a robust and miniaturisable bio-photonic platform with direct implications for quantum light sensing, neuromorphic bioelectronics, and next-generation artificial retinal interfaces.
]]></description>
<dc:creator><![CDATA[ Cardace, I., Dominici, L., Ardizzone, V., Cola, A., Fieramosca, A., Nobile, C., Polticelli, F., Bruni, F., De Giorgi, M., Ballarini, D., Gigli, G., De Marco, L., Sanvitto, D. ]]></dc:creator>
<dc:date>2026-05-31</dc:date>
<dc:identifier>doi:10.64898/2026.05.29.728716</dc:identifier>
<dc:title><![CDATA[Electrically programmable picoscale phototransduction of a newly discovered microbial rhodopsin]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-31</prism:publicationDate>
<prism:section></prism:section>
</item>
<item rdf:about="https://www.biorxiv.org/content/10.64898/2026.05.28.728512v1?rss=1">
<title>
<![CDATA[
Protonated Structure of EmrE Reveals C-terminal Tail Gating Mechanism 
]]>
</title>
<link>
https://www.biorxiv.org/content/10.64898/2026.05.28.728512v1?rss=1
</link>
<description><![CDATA[
The multidrug efflux pump EmrE is one of the smallest known active transporters and has become a model system for studying multidrug recognition and transport. While recent high-resolution structures have illuminated its dynamic substrate binding pocket, the conformations of its interhelical loops and C-terminal tail, regions critical for controlling proton coupling and gating, remain poorly characterized. Here, we report the high-resolution structure of protonated S64V EmrE determined using solution and solid-state NMR data. This new structural model shows the C-terminal tail occluding the open face of the transport pore, providing a structural basis for how EmrE minimizes proton leak in the absence of substrate. These findings support growing evidence that relatively simple model transporters must leverage an occluded state during alternating access to avoid physiologically unfavorable proton leak.
]]></description>
<dc:creator><![CDATA[ Hiett-Borcik, A. B., Harding, B. D., Brousseau, M., Warmuth, O., Borcik, C. G., Wu, C., Uhlemann, E.-M., Cornilescu, C. C., Reichert, G. E., Henzler-Wildman, K. ]]></dc:creator>
<dc:date>2026-05-30</dc:date>
<dc:identifier>doi:10.64898/2026.05.28.728512</dc:identifier>
<dc:title><![CDATA[Protonated Structure of EmrE Reveals C-terminal Tail Gating Mechanism]]></dc:title>
<dc:publisher>Cold Spring Harbor Laboratory</dc:publisher>
<prism:publicationDate>2026-05-30</prism:publicationDate>
<prism:section></prism:section>
</item>
</rdf:RDF>
