bims-enlima 2025-12-28 papers

bims-enlima

Biomed News

on Engineered living materials

Issue of 2025–12–28
forty-one papers selected by
Rahul Kumar, Tallinna Tehnikaülikool

Biological Complexity-Inspired Engineering of Tough and Anisotropic Protein-Based Materials for Adaptive Sensing.
Melt Densification Enables Fracture-Resistant Blend Hydrogels.
Polymeric Artificial Cells: from Interfacial Membranization to Cytomimetic Architecture Engineering.
Extended networks.
Active learning-guided optimization of cell-free biosensors for lead testing in drinking water.
Light-Based 3D Printing of Polyesters: From Synthesis to Fabrication.
A stationary phase-specific bacterial green light sensor for enhancing metabolite production.
3D Printing-Assisted Interpenetrating-Phase Composite Implant Materials Integrating Structural-Functional Properties.
Ultra-Fast Isothermal Formation of DNA Nanostructures in Culture Media: Application to In Situ Assembly of DNA Origami With Living Cells.
Low-Cost Open Platform Digital Light Printer (OP-DLP) for 96-Well Format Hydrogel Printing and Localized Light-Activation.
Percolation by Brush Architecture: A Pathway toward Soft Electronics.
Piezoelectric Film-Based Bacterial Disinfection by Mechanical Agitation.
Bioinspired Flexible Tactile Sensors for Smart Soft Robotics.
Arabinose-Inducible Univariant Control System (AUCS) for Microbial Production of Proteins, Enzymes, and Metabolites.
Sensing the shape of a surface by tightly surface-bound filaments.
Physics-Informed Graph Neural Networks for Predicting Deformation in Disordered Fibrous Materials.
Alginate Hydrogels with Tunable Degradation.
Protocol to identify mechanosensitive nuclear proteins using tunable actomyosin contractility and proximity biotinylation in mammalian cells.
Traceless Regulation of Genetic Circuitry.
Synthetic Control of Implanted Engineered Liver Tissue Growth.
Self-assembling protein cages: from coiled-coil module to machine learning-driven de novo design of next-generation biomaterials.
DNA-guided 3D pathway reconfiguration of composite nanopores for ion transport regulation.
Engineering artificial biosynthetic pathway enables simultaneous production and in-situ bio-dyeing of indigoids for textiles.
Intracellularly coupled oscillators for synthetic biology.
tRNA-Mediated Plasmid Stabilization for Antibiotic-Free Applications in Escherichia coli.
Protocol for marker-free genome editing in Saccharomyces cerevisiae using universal donor templates and multiplexed CRISPR-Cas9.
Photocontrolled trimethoprim PROTACs targeting the eDHFR protein tag.
DNA-based hydrogels: a promising material for future energy storage applications.
Getting gene expression "just right".
Protein force spectroscopy using magnetic tweezers: slow and steady wins the race?
Integrating Vibrio natriegens for Photon Manipulation in Living Lighting Devices.
Computational design of allulose-responsive biosensor toolbox for auto-inducible protein expression and CRISPRi mediated dynamic metabolic regulation.
Responsive Emulsions Enabled by Supramolecular Polymers for Recyclable Biocatalysis.
Multiplex mapping of protein-protein interaction interfaces.
A Simple and Versatile Cell-Free Expression Method for Producing Secondary Metabolites.
Maintenance of cytoplasmic and membrane densities shapes cellular geometry in Escherichia coli.
Closed-loop optogenetic control of cell biology enables outcome-driven microscopy.
Designing synthetic regulatory elements using the generative AI framework DNA-Diffusion.
Design of a minimal, allosteric, and ATPase-like machine using mechanical linkages.
Genetic Code Expansion in Probiotics Enables the Secretion of Covalent Protein Drugs in Mice.
High-Throughput Detection of Cyanobacterial Form I Rubisco Assembly.

Nano Lett. 2025 Dec 20.

Biological Complexity-Inspired Engineering of Tough and Anisotropic Protein-Based Materials for Adaptive Sensing.

Zhe Lu, Zhenhao Zhu, Hao Lu, YaoLong Zhi, Hong Hu, You Yu.

  Biological systems inspire the design of high-performance biomimetic materials, yet replicating their synergistic interactions and hierarchical structures remains challenging. Here, we present an orthogonal photochemistry-mediated strategy for one-step fabrication of muscle-inspired protein materials. This approach integrates covalent, electrostatic, and hydrogen-bonding interactions to form robust multinetwork architectures within hierarchically organized protein matrices. Prestretching enhances molecular alignment, yielding anisotropic materials with a factor of 3.0, tensile strengths up to 300 MPa, toughness over 22 MJ m-3, and a fatigue threshold of 760 J m-2─surpassing natural proteins such as wool, cotton, and silk. The rapid (∼20 s), precisely controllable process supports scalable 3D manufacturing of continuous fibers exceeding 10 m. Beyond mechanical robustness, the materials dynamically respond to force, humidity, and pH, mimicking biological tissues. As a proof of concept, the fibers function as artificial muscles and flexible capacitive sensors, highlighting their potential for advanced applications in biomaterials, bioengineering, and soft electronics.

Keywords:  3D printing; anisotropy; photochemistry; protein materials; tough hydrogels

DOI:  https://doi.org/10.1021/acs.nanolett.5c04826
Adv Mater. 2025 Dec 24. e17395

Melt Densification Enables Fracture-Resistant Blend Hydrogels.

Xunan Hou, Zichun Zhu, Yuting Wen, Yixin Zhang, Chitinart Thedrattanawong, Daria V Andreeva, Jun Li, Chaobin He.

  Hydrogels and elastomers are integral components in biomedical and electronics devices, but their toughness and crack resistance are often unsatisfactory for load-bearing applications. Synthetic polymer networks predominantly rely on solution fabrication, which compromises the ultimate mechanical properties. This work presents a universal melt crosslinking strategy, which densifies entanglements well beyond solvated conditions. When deformed, mutually entangled dissimilar chains stiffen the gels, while sparse crosslinks amplify fracture resistance. At water contents up to 83%, the resultant hydrogels demonstrate over 2 orders increase in mechanical properties, including moduli (1.3-35 MPa), toughness (0.7-24.5 kJ/m2), and fatigue thresholds (1.2-3.3 kJ/m2), tunable in a wide range beyond existing hydrogels. Furthermore, the hydrogels show high optical clarity (>96%), oxygen permeability (Dk/t > 40), and anti-fouling properties (<0.6 µg cm-2). This generalizable strategy could guide the design of tough functional soft materials in fields such as healthcare and smart electronics.

Keywords:  antifouling; hydrogels; polymers; resistance; toughening

DOI:  https://doi.org/10.1002/adma.202517395
Biomacromolecules. 2025 Dec 23.

Polymeric Artificial Cells: from Interfacial Membranization to Cytomimetic Architecture Engineering.

Hao Han, Siyu Song, Yubin Pu, Tsvetomir Ivanov, Shoupeng Cao.

Artificial cells emulating the structure and function of living systems have attracted tremendous research attention. By integrating concepts and techniques from chemistry, materials science, and biochemistry, researchers have assembled functional building modules into advanced materials capable of exhibiting life-like behaviors. Recent advances have led to the creation of synthetic cells that mimic key characteristics of living cells. Polymer-based systems attract enormous interest due to their chemical versatility, robustness, and programmability. These attributes allow the construction of artificial cells with precise modulation of physicochemical properties, architecture and functionality. Representative polymeric artificial cells have demonstrated essential hallmarks of life, including membranization, integration of suborganelles, and formation of cytoskeletal frameworks. In this Review, we highlight recent advances in the design, assembly, and functionalization of polymer-based artificial cells, emphasizing how the intrinsic tunability and multifunctionality of polymeric materials enable the recreation and extension of life-like structures, dynamic behaviors, and biological functions.

DOI: https://doi.org/10.1021/acs.biomac.5c02327
Nat Mater. 2025 Dec 24.

Extended networks.

DOI: https://doi.org/10.1038/s41563-025-02466-6
Nat Commun. 2025 Dec 20.

Active learning-guided optimization of cell-free biosensors for lead testing in drinking water.

Brenda M Wang, Nicole Chiang, Holly M Ekas, Dylan M Brown, Garrett Dildine, Tyler J Lucci, Siyuan Feng, Vanessa Bly, Jean-François Gaillard, Julius B Lucks, Ashty S Karim, Diwakar Shukla, Michael C Jewett.

Point-of-use diagnostics based on allosteric transcription factors (aTFs) are promising tools for environmental monitoring and human health. However, biosensors relying on natural aTFs rarely exhibit the sensitivity and selectivity needed for real-world applications, and traditional directed evolution struggles to optimize multiple biosensor properties at once. To overcome these challenges, we develop a multi-objective, machine learning (ML)-guided cell-free gene expression workflow for engineering aTF-based biosensors. Our approach rapidly generates high-quality sequence-to-function data, which we transform into an augmented paired dataset to train an ML model using directional labels that capture how aTF mutations alter performance. We apply our workflow to engineer the aTF PbrR as a point-of-use diagnostic for lead contamination in water. We tune the sensitivity of PbrR to sense at the U.S. Environmental Protection Agency (EPA) action level for lead and modify the selectivity away from zinc, a common metal found in water supplies. Finally, we show that the engineered PbrR functions in freeze-dried cell-free reactions, enabling a diagnostic capable of detecting lead in drinking water down to ~5.7 ppb. Our ML-driven, multi-objective framework powered by directional tokens can generalize to other biosensors and proteins, accelerating the development of synthetic biology tools for biotechnology applications.

DOI: https://doi.org/10.1038/s41467-025-66964-6
Chem Rev. 2025 Dec 23.

Light-Based 3D Printing of Polyesters: From Synthesis to Fabrication.

Quinten Thijssen, Astrid Quaak, Bart Bijleveld, Bo Li, Lenny Van Daele, Andreas Heise, Sandra Van Vlierberghe.

Polyesters represent a versatile class of materials whose biodegradability, biocompatibility, mechanical tunability, and broad chemical design space have made them valuable across a wide range of application areas, including tissue engineering, biomedical engineering, sustainable manufacturing, and soft robotics. Light-based 3D printing has further expanded their potential by enabling precise spatial control across nano- to macroscales, supporting the fabrication of resorbable implants, drug-delivery systems, microneedle arrays, and stimuli-responsive materials. This review discusses the essential steps toward light-based 3D printing of polyesters from synthetic strategies for producing these materials to functionalization methods that render them suitable for light-based 3D printing. Particular attention is given to the synthetic origin of the polyester, the way photoreactive groups are introduced and organized within the network, and how the formulation of the resulting photoresin together govern the ultimate photoreactivity, degradation behavior, print resolution, and mechanical performance. Advantages and limitations of current photochemical approaches are discussed across different light-based 3D printing technologies. With continuing advancements in manufacturing, the field of light-based 3D printing of polyesters shows substantial promise, poised to redefine material design, and influence a broad range of future technologies.

DOI: https://doi.org/10.1021/acs.chemrev.5c00611
Nat Commun. 2025 Dec 24.

A stationary phase-specific bacterial green light sensor for enhancing metabolite production.

John T Lazar, Daniel J Haller, Abbas Ghaddar, Jae J Kim, Kevin Yang, Sebastián M Castillo-Hair, Andrew R Gilmour, Ross Thyer, Jeffrey J Tabor.

Genetically-encoded sensors are used to control protein and metabolite production in bacterial fermentations. However, these sensors are generally optimized for exponential growth rather than stationary phase where production occurs. Here, we find that our previously engineered E. coli green light sensor CcaSR, which functions robustly in exponential phase, fails in stationary phase due to spontaneous loss of an engineered chromophore biosynthetic pathway and accumulation of CcaS and CcaR. We optimize the genetic context and expression determinants of each component, resulting in a stable system named CcaSRstat that imposes little metabolic burden, exhibits low leakiness and an 80-fold green light response, and functions exclusively in stationary phase. We combine CcaSRstat-driven enzyme expression with varied static and periodic illumination patterns to achieve high titers of the industrially-relevant phenylpropanoid p-Coumaric acid and demonstrate that these optimizations scale to benchtop bioreactor conditions. Finally, we use CcaSRstat to optimize the expression level of a co-transcribed multi-enzyme metabolic pathway encoding production of plant-derived betaxanthin family pigments. Stationary phase-optimized bacterial sensors should enhance fermentation productivity by enabling rapid interrogation of the impact of enzyme expression level and induction dynamics.

DOI: https://doi.org/10.1038/s41467-025-67829-8
Adv Mater. 2025 Dec 26. e14145

3D Printing-Assisted Interpenetrating-Phase Composite Implant Materials Integrating Structural-Functional Properties.

Jiaxing Huo, Yanyan Liu, Zengqian Liu, Ning Wang, Baohong Zhao, Jiantao Li, Lili Tan, Yufeng Zheng, Zhefeng Zhang, Robert O Ritchie, Qiang Wang.

  Implant materials play a pivotal role in bone repair; however, existing materials face considerable challenges in simultaneously achieving adequate mechanical properties and biological functions. In this perspective, drawing inspiration from nature and leveraging 3D printing technologies, we propose a new strategy to achieve structural-functional integration through the development of bicontinuous interpenetrating-phase composite implant materials. These materials are fabricated by infiltrating one constituent into 3D-printed porous scaffolds of another, and demonstrate two potential degradation pathways after implantation - selectively partial degradation and sequentially complete degradation - depending on the types of constituents and their combinations. We elucidate the associated degradation behaviors, regulatory strategies, and the resulting biological functions, and analyze their underlying cellular and molecular mechanisms. Moreover, targeted functional integration and delivery can be realized by infiltrating hydrogels loaded with functional agents into 3D-printed scaffolds. The mechanical and functional properties of these materials can be deliberately modulated by selecting appropriate constituents and by designing and regulating the interpenetrating-phase structures. We further examine the challenges faced by these materials and outline prospective directions for future research. Distinct from conventional single-component materials, 3D printing-assisted composite implant materials hold significant promise for achieving structural-functional integration, thereby offering new opportunities to enhance bone repair efficacy.

Keywords:  3D printing; bone repair; implant materials; interpenetrating‐phase composites; structural‐functional integration

DOI:  https://doi.org/10.1002/adma.202514145
Small. 2025 Dec 23. e09401

Ultra-Fast Isothermal Formation of DNA Nanostructures in Culture Media: Application to In Situ Assembly of DNA Origami With Living Cells.

Laura Bourdon, Gerrit David Wilkens, Samy Dehissi, Salammbô Hotte, Sergii Rudiuk, Mathieu Morel, Ayako Yamada, Gaëtan Bellot, Damien Baigl.

  Synthetic DNA strands are programmable and biocompatible building blocks that can be combined through hybridization to form user-defined nanostructures, but their assembly traditionally requires cell-incompatible conditions, imposing a lengthy ex situ fabrication step before any application with living matter. Here we demonstrate for the first time that 2D and 3D DNA origami structures can isothermally self-assemble at 37°C within minutes, directly in cell culture media, both in the absence and in the presence of living cells. Scaffold-free structures of extended dimensions, such as micrometer-long DNA nanotubes, can also self-assemble when the system is given more time to evolve. With human cell lines, 2D and 3D origami structures in situ self-assemble in 5 to 15 min, and remain stable for about 24 h and up to 3 days when actin monomers are added. Similar self-assembly performance is observed in the presence of more complex tissue-like systems, such as human induced pluripotent stem cells evolving into cerebral organoids. This ultra-fast, life-compatible self-assembly method drastically simplifies the fabrication of complex DNA nanostructures and enables the creation of in situ self-assembling nanomachines for direct and adaptive interactions with living cells.

Keywords:  DNA nanotechnology; bio‐interface; brain organoid; nanomachine; self‐assembly

DOI:  https://doi.org/10.1002/smll.202509401
ACS Biomater Sci Eng. 2025 Dec 26.

Low-Cost Open Platform Digital Light Printer (OP-DLP) for 96-Well Format Hydrogel Printing and Localized Light-Activation.

Katelyn Mathis, Afia Ibnat Kohon, Natanael Monroy, Lamees Abu-Suleiman, Amanda Yang, Brian Meckes.

  There is growing demand for high-throughput light-based surface processing methods for applications such as printing hydrogels, controlling cell circuits with light, or activating materials on demand. However, existing devices often fall short for multiwell plate use, require complex synthesis steps, or lack flexibility for general research needs, usually because they are designed for specific tasks. Here, an open-platform digital light printer (OP-DLP) is introduced for easy synthesis of two-dimensional (2D) hydrogels and spatial activation of biomolecules. The device is controlled via a LabVIEW interface that manages printing settings and planar corrections. Importantly, its open platform design enables the use of different wavelengths and compatibility with various printing vessels. Its utility is demonstrated by hydrogel printing and spatial activation of DNA. Specifically, OP-DLP can produce hydrogel layers of precise thickness in a 96-well format with consistent results across the plate. Additionally, OP-DLP can form 2D gels with specific shapes in different wells, allowing modification of ink composition. Its spatial activation capability is demonstrated by the localized de-caging of photocaged DNA on a surface.

Keywords:  biomaterials; devices; hydrogel printing; light-activation

DOI:  https://doi.org/10.1021/acsbiomaterials.5c01894
ACS Appl Mater Interfaces. 2025 Dec 26.

Percolation by Brush Architecture: A Pathway toward Soft Electronics.

Josiah H Marshall, Zilu Wang, Liang Yan, Wei You, Akmal Z Umarov, Ioannis Moutsios, Dimitri A Ivanov, Andrey V Dobrynin, Sergei S Sheiko.

  Achieving simultaneous tissue-mimetic mechanical properties and electrical conductivity within a single molecular system remains a major challenge. Typically, the design of stretchable and flexible electronic materials requires a trade-off between mechanical compliance and electronic performance. Herein, we employ a computationally driven materials design strategy to synthesize polydimethylsiloxane bottlebrush graft copolymers with precisely controlled fractions of poly(3-hexylthiophene) (P3HT), where the grafts serve both as physical cross-links between bottlebrush strands and as conductive elements. Thin films cast from solution reveal percolation of P3HT needle-like crystals, corroborated by transmission electron microscopy, small-angle X-ray scattering, and computer simulations. These films exhibit a distinctive combination of properties, including a low elastic modulus (∼1-100 kPa) and an electrical conductivity of up to ∼10-2 S/cm. Materials that simultaneously combine tissue-like mechanics with electronic functionality hold strong potential for wearable and implantable devices.

Keywords:  P3HT crystallization; bottlebrush polymers; conductive elastomers; polymer networks; tissue-mimetic mechanical properties

DOI:  https://doi.org/10.1021/acsami.5c19739
ACS Appl Mater Interfaces. 2025 Dec 24.

Piezoelectric Film-Based Bacterial Disinfection by Mechanical Agitation.

Debiprasad Roy, Buddhadev Mukherjee, Swapnendu Deb, Saikat Kumar Pandit, Jayanta Dolai, Soumya Dhang, Subhajit Saha, Arindam Banerjee, Nikhil R Jana.

  The activation of piezoelectric materials by using mechanical pressure offers a promising approach for both environmental remediation and biomedical therapeutics. Here, we report BaTiO3 nanorod-embedded polymer film/membrane that can be used for bacterial disinfection via mechanical stress-induced chemical reaction and reactive oxygen species (ROS) generation. Incorporating 6 wt % BaTiO3 nanorods into the polyvinylidene fluoride/poly l-lactic acid matrix led to the maximum enhancement (with a piezoelectric constant of 66 pm/V) of both the piezoelectric properties and the mechanochemical ROS generation. These films are utilized for sono-piezocatalytic disinfection of both Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus (>80% inactivation) and observed that the piezoelectric properties of films correlate with ROS generation property and antibacterial activity. Further, a piezoelectric reactor is developed for water flow-based treatment of contaminated water by integrating these piezoelectric membranes. Results show that water flow-based mechanical stress can disinfect E. coli, S. aureus, and degrade chemical pollutants. Proposed materials and approaches can be extended for wastewater treatment and related applications.

Keywords:  BaTiO3 nanoparticle; antibacterial materials; mechanochemistry; piezocatalysis; piezoelectric; reactive oxygen species; ultrasound

DOI:  https://doi.org/10.1021/acsami.5c16510
ACS Appl Mater Interfaces. 2025 Dec 22.

Bioinspired Flexible Tactile Sensors for Smart Soft Robotics.

Zhongbao Wang, Yi Tang, Pengyu Yao, Yixin Chen, Jingyue Luo, Wenhao Xue, Yuan Ma, Jing Wang.

  This review delves into the cutting-edge advancements in the bioinspired flexible tactile sensors and their transformative role in enabling smart soft robots with embodied intelligence. Drawing inspiration from the multifunctionality of human skin, tactile sensors are desired to include similar mechanoreceptor, proprioception, and environmental responsiveness. This review begins by outlining fundamental design principles that mimic the hierarchical structure and distributed sensing networks of human skin. The biomimetic design and sensing principles of different flexible tactile sensors are then explained and compared, including pressure, temperature, and strain sensors. The state-of-the-art manufacturing methods, including direct ink writing, fused deposition modeling, digital light processing and material jet printing, are also introduced. By summarizing typical applications of these tactile sensors in smart soft robots, delicate object manipulation, human-robot collaboration, medical prosthetics, and adaptive locomotion are primarily discussed. Finally, it is promising to integrate innovations in fatigue-resistant elastomers, nanometer-scale 3-dimensional manufacturing, and artificial intelligence as potential elements to create next-generation of tactile sensors in the near future. By bridging biomimetic design, soft materials and robotics, this review aims to equip researchers and engineers with the knowledge to develop the tactile systems that push the boundaries of autonomy, safety, and interaction in soft robotics.

Keywords:  bioinspired engineering; electronic skin; flexible electronics; soft robotics; tactile sensors

DOI:  https://doi.org/10.1021/acsami.5c16200
ACS Synth Biol. 2025 Dec 21.

Arabinose-Inducible Univariant Control System (AUCS) for Microbial Production of Proteins, Enzymes, and Metabolites.

Congqiang Zhang, Sudha Shukal.

  Precise control of gene expression is essential to synthetic biology and metabolic engineering, particularly for microbial production. The widely used IPTG-inducible T7lac promoter (PT7lac) offers strong expression but suffers from metabolic burden, inclusion body formation, and induction heterogeneity. Conversely, the arabinose-inducible araBAD promoter (PBAD) provides tight regulations but yields modest expression levels, is incompatible with glucose media, and requires high inducer concentrations (20-100 mM). We introduce the arabinose-inducible univariant control system (AUCS), a robust, tightly regulated, and low-cost expression platform designed to combine the strengths of PT7lac and PBAD while overcoming their drawbacks. AUCS eliminates carbon catabolite repression and minimizes induction heterogeneity via the constitutive expression of the arabinose transporter AraE. Disruption of the arabinose catabolism enables maximal protein output with only 3 μM l-arabinose, orders of magnitude lower than PT7lac and PBAD systems, achieving a >99% reduction in inducer cost. Leveraging a customized promoter library (PTA1-3), AUCS enables the precise, high-yield expression of single proteins, multienzyme operons, and complex biosynthetic pathways (>10 genes). Benchmarked against PT7lac, AUCS achieved comparable or superior yields of proteins (the egg-white protein ovalbumin), enzymes (terpene synthases, carotenoid cleavage dioxygenases), and secondary metabolites (linalool, nerolidol, and sclareol) while maintaining outstanding reproducibility and stability over 36 generations. AUCS represents a powerful advancement for precision fermentation, enabling sustainable and cost-effective production of high-value biomolecules and substantially reducing the environmental footprint of chemical manufacturing.

Keywords:  Synthetic biology; arabinose; gene regulation; metabolic engineering; microbial control system; regulatory element

DOI:  https://doi.org/10.1021/acssynbio.5c00602
Proc Natl Acad Sci U S A. 2025 Dec 30. 122(52): e2526131122

Sensing the shape of a surface by tightly surface-bound filaments.

Handuo Shi, Jeffrey Nguyen, Jordan Alexander Huang, Zemer Gitai, Joshua Shaevitz, Benjamin P Bratton, Ajay Gopinathan, Gregory Grason, Kerwyn Casey Huang.

  Understanding the mechanisms that dictate the localization of cytoskeletal filaments is crucial for elucidating cell shape regulation in prokaryotes. The actin homolog MreB plays a pivotal role in maintaining the shape of many rod-shaped bacteria such as Escherichia coli by directing cell-wall synthesis according to local curvature cues. However, the basis of MreB's curvature-dependent localization has remained elusive. Here, we develop a biophysical model for the energetics of a filament binding to a surface that integrates the complex interplay between filament twist and bending and the two-dimensional surface geometry. Our model predicts that the spatial localization of a filament like MreB with substantial intrinsic twist is governed by both the mean and Gaussian curvatures of the cell envelope, which strongly covary in rod-shaped cells. Using molecular dynamics simulations to estimate the mechanical properties of MreB filaments, we show that their thermodynamic preference for regions with lower mean and Gaussian curvatures matches experimental observations for physiologically relevant filament lengths of ~50 nm. We find that the experimentally measured statistical curvature preference is maintained in the absence of filament motion and after a cycle of depolymerization, repolymerization, and membrane rebinding, indicating that equilibrium energetics can explain MreB localization. These findings provide critical insights into the physical principles underlying cytoskeletal filament localization and suggest design principles for synthetic shape-sensing nanomaterials.

Keywords:  Gaussian curvature; MreB; curvature enrichment; cytoskeletal filaments; intracellular localization

DOI:  https://doi.org/10.1073/pnas.2526131122
Nano Lett. 2025 Dec 26.

Physics-Informed Graph Neural Networks for Predicting Deformation in Disordered Fibrous Materials.

Shuo Yang, Yunhao Yang, Chen Huang, Runnan Bai, Leitao Cao, Jing Ren, Shengjie Ling.

  Disordered fibrous networks play vital mechanical roles but are difficult to model due to their sparse connectivity and complex nonaffine deformation. We introduce a physics-informed, graph-learning-based Network Mechanics Prediction (GNMP) that predicts deformation and stress-strain responses of 2D semiflexible networks with reliable accuracy and >10× efficiency gains over molecular dynamics. GNMP integrates graph-attention message passing, multiscale physical embeddings, and a bond-length-guided scheduler to capture key nonaffine rearrangements. Experiments on Poisson-ratio-aligned 3D-printed networks show consistent deformation motifs, descriptor trends, and comparable log-area changes (∼0.46 vs ∼0.24) while reducing the geometry-inference time to ∼0.13 s per step. GNMP offers a generalizable route for rapid, topology-aware mechanical prediction in fibrous network materials and supports the accelerated design of biomimetic soft tissues, flexible conductors, and related network-based systems.

Keywords:  Computational materials design; Disordered fibrous networks; Nonaffine deformation; Physics-informed graph neural networks; Stress−strain prediction

DOI:  https://doi.org/10.1021/acs.nanolett.5c04895
Macromol Rapid Commun. 2025 Dec 22. e00348

Alginate Hydrogels with Tunable Degradation.

Haoyi Hou, Alison M Lanzi, Yunpeng Feng, Raavi, Arturo J Vegas.

  Hydrogels have found utility as both tissue scaffolds and cell delivery vehicles. As tissue engineering scaffolds, they provide physical and biochemical cues that guide tissue remodeling processes. Degradation of the gel over time is a desirable material property that allows for the complete integration of imbued cells with the native tissue. Here, novel alginate-based hydrogels with tunable degradability were formulated by synthesizing diblock polymers with a degradable domain. Alginate and poly(D, L-lactic acid) (PLA) of varying molecular weights were end-modified using bio-orthogonal chemistry and linked covalently to form alginate-b-PLA diblock polymers. These polymers retained both the ionic-cross-linking properties of alginate and the hydrolytic degradation properties of PLA. The hydrogel degradation rate was determined by the size of the constituent domains as well as the diblock content in blended gels and enabled the tunable, temporal release of encapsulated cells. This material platform is useful where tunable degradation is advantageous, such as in regenerative medicine or drug delivery.

Keywords:  alginate hydrogel; biorthogonal conjugation; cell delivery; degradable alginates; tissue engineering; tunable cell deposition; tunable degradability

DOI:  https://doi.org/10.1002/marc.202500348
STAR Protoc. 2025 Dec 24. pii: S2666-1667(25)00694-X. [Epub ahead of print]7(1): 104288

Protocol to identify mechanosensitive nuclear proteins using tunable actomyosin contractility and proximity biotinylation in mammalian cells.

Pei-Li Tseng, Weiwei Sun, Jiawei Li, Mark O Collins, Kai S Erdmann.

  Mechanical forces influence a range of cellular behaviors; however, how these forces are sensed and converted into biochemical changes remains incompletely understood. A key aspect of mechanotransduction is the regulation of subcellular protein localization. Here, we present a protocol describing the engineering of cell lines with tunable actomyosin contractility combined with a proximity biotinylation strategy confined to the nucleus followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. This approach allows the identification of proteins whose nuclear localization is controlled by changes of actomyosin contractility. For complete details on the use and execution of this protocol, please refer to Tseng et al.1.

Keywords:  Biophysics; Cell biology; Molecular biology

DOI:  https://doi.org/10.1016/j.xpro.2025.104288
Adv Sci (Weinh). 2025 Dec 27. e19848

Traceless Regulation of Genetic Circuitry.

Gokberk Unal, Martin Fussenegger.

  The advent of synthetic biology, enabling the construction of synthetic genetic circuitry with designed functionality, has has a revolutionary impact on medicine, agriculture, sustainable energy, and the industrial production of high-value compounds over the last few decades. Gene switches have an indispensable role as regulators of such systems. Despite the early introduction of chemically inducible switches to regulate genetic circuitry, 'traceless' physical cues (e.g., light, heat, sound, magnetism, electricity, and mechanical force) can provide greater specificity, higher spatiotemporal resolution, more flexible switching patterns, and better compatibility with bioelectronic interfaces, which is of particular significance given the rise of electrogenetics. Indeed, traceless gene switches are on a path to become universal biological control ports interfacing physiology with the electronic world. In this review, we discuss the impact, challenges, and prospects of physically inducible, traceless gene switches in the context of recent cutting-edge applications.

Keywords:  cell circuit; gene switch; synthetic biology; traceless

DOI:  https://doi.org/10.1002/advs.202519848
bioRxiv. 2025 Dec 12. pii: 2025.12.10.693527. [Epub ahead of print]

Synthetic Control of Implanted Engineered Liver Tissue Growth.

Amy E Stoddard, Vardhman Kumar, Constantine N Tzouanas, Veronica Hui, Jeffrey Li, Anisha Jain, Alanna Farrell, Sangeeta N Bhatia, Christopher S Chen.

Despite the promise of engineered tissue implants for the treatment of organ failure, scaling of these constructs to sizes of therapeutic relevance remains a barrier to clinical translation. Here, we propose a strategy to circumvent this limitation: to instead implant a small-scale construct and then induce it to grow in situ after its engraftment into a host. Using engineered liver tissue as a proof-of-concept application, we integrated synthetic biology and tissue engineering tools to build liver tissues that can be expanded on-demand after implantation in vivo . To achieve this goal, we first identified the combination of YAP and growth factor signaling as sufficient to drive human hepatocyte proliferation in dense, 3D engineered tissues. We then engineered control of these signaling axes using synthetic biology tools to drive human liver tissue expansion both in vitro and in vivo . As such, this work establishes a genetic strategy for generating large organ implants through bioengineered, on-demand outgrowth using synthetic triggers (BOOST).
Teaser: Presenting bioengineered on-demand outgrowth via synthetic biology triggering (BOOST) for in situ solid cell therapy scale up.

DOI: https://doi.org/10.64898/2025.12.10.693527
Mater Adv. 2025 Dec 23.

Self-assembling protein cages: from coiled-coil module to machine learning-driven de novo design of next-generation biomaterials.

Arvind Kumar Gupta, Hana Esih, Helena Gradišar, Roman Jerala.

The rational design of self-assembling protein nanocages holds great promise for synthetic biology, biotechnology and biomedical applications. Protein nanocages are well-defined nanoparticles with an inner cavity formed by self-assembly of repetitive protein building blocks. These cavities can be tailored to encapsulate and protect cargo molecules such as drugs, enzymes, or imaging agents. The ability to design de novo protein cages has recently been revolutionized by new concepts of modular protein design, computational design of interacting surfaces and machine learning-based generative protein design. Protein cages can be designed in diverse architectures and sizes, and their assembly and disassembly can be regulated by chemical, biological, and physical signals. Here, we focus on the review of engineering strategies for the designed protein cages based on coiled coils or other modular protein domains, their functionalization and opportunities of customized engineered protein cages.

DOI: https://doi.org/10.1039/d5ma00792e
J Nanobiotechnology. 2025 Dec 24. 23(1): 788

DNA-guided 3D pathway reconfiguration of composite nanopores for ion transport regulation.

Jiarong Guo, Tao Gao, Ying Ma, Jinwen Zhu, Jiayue Shi, Baoguo Wang, Peng Miao.

  Ion current rectification (ICR) is essential for understanding analyte-driven nanofluid transport within nanopores. However, the rapid flow rates and limited reaction times in this process can impede electrochemical reactions at electrode interfaces, which consequently lead to electrical noise challenges. Here, we propose a simple strategy to enhance the ICR effect and reduce noise interference through the construction of bio/solid composite pores. The composite pores comprise θ-shaped glass pores that have been sequentially modified with 3-glycidyloxypropyltrimethoxysilane (GLYMO), succinic anhydride (SA) and single-stranded DNA. Three-dimensional (3D) biochannels formed from acrylamide-DNA hydrogels are integrated within glass pores. High-density 3D channels and a highly viscous environment can decelerate analyte traversal, thereby establishing a stable reaction environment. Concurrently, acrylamide-DNA hydrogels modulate pore size through cascade reactions triggered by analytes, thereby altering the transport pathways of ion and affecting ICR. Furthermore, we have developed a sensor featuring adjustable ion transport pathways based on this technology. The detection range can be readily expanded to include nucleic acids, proteins, glycans and a multitude of biomolecules through modifying different aptamers.

Keywords:  Composite nanopore; DNA hydrogel; Electrochemical sensing; Ion current rectification; Strand displacement

DOI:  https://doi.org/10.1186/s12951-025-03849-2
Nat Commun. 2025 Dec 26.

Engineering artificial biosynthetic pathway enables simultaneous production and in-situ bio-dyeing of indigoids for textiles.

Qi Na, Xiaowang Zhang, Linna Long, Siyu Qi, Zhiyu Zhang, Ruoyu Zhi, Zhuotao Tan, Hanjie Ying, Chenjie Zhu.

The textile dyeing industry has shaped modern aesthetics through vibrant coloration technologies. Indigoid dyes embody this potential but are hindered by the hazardous chemo-synthesis and water-intensive chemo-dyeing processes. While biosynthesis and bio-dyeing efforts offer alternatives, disjointed synthesis-dyeing steps lead to unsustainable water and energy demands, and complex biosynthetic pathways cause low atom economy and inefficiency. Here, we report a one-pot one-step approach that unifies biosynthesis and bio-dyeing of indigoids to drastically reduce water usage in the dyeing process. By combining alcohol dehydrogenase with flavin-containing monooxygenase, we construct an artificial redox-neutral cascade biosynthetic pathway to produce bluish-violet indigoid dyes, which significantly enhances atom economy and achieves high production efficiency. Various natural and synthetic fabrics are dyed in-situ with the biosynthetic indigoids utilizing recombinant E. coli whole cells. Crucially, this system dyes synthetic nylon fibers with the historic purple hue.

DOI: https://doi.org/10.1038/s41467-025-67935-7
Nat Commun. 2025 Dec 21.

Intracellularly coupled oscillators for synthetic biology.

Gábor Holló, Jung Hun Park, Rose A Evard, Yolanda Schaerli.

Synthetic biology aims to engineer or re-engineer living systems. To achieve increasingly complex functionalities, it is beneficial to use higher-level building blocks. In this study, we focus on oscillators as such building blocks, propose oscillator-based circuit designs and model the interactions of intracellularly coupled oscillators. We classify these oscillators on the basis of coupling strength: independent, weakly or strongly, and deeply coupled. We predict a wide range of dynamic behaviors to arise in these systems, such as the beat phenomenon, amplitude and frequency modulation, period doubling, higher-period oscillations, chaos, resonance, and synchronization, with the aim of guiding future experimental work in bacterial synthetic biology. Finally, we outline potential applications, including oscillator-based computing that integrates processing and memory functions, offering multistate and nonlinear processing capabilities.

DOI: https://doi.org/10.1038/s41467-025-67893-0
ACS Synth Biol. 2025 Dec 23.

tRNA-Mediated Plasmid Stabilization for Antibiotic-Free Applications in Escherichia coli.

Ana G V Sepulchro, Andreas B Bertelsen, Ivan Schlembach, Marta R Montané, Suresh Sudarsan, Morten H H No̷rholm, Viji Kandasamy.

  Plasmids are essential tools in molecular biology and biotechnology. In research laboratories, it is common to use antibiotic selection markers to ensure that plasmids are stably maintained in a cellular population. However, the use of antibiotics poses a significant challenge in the industrial scale-up process due to the high cost and the risk of spreading resistance. Therefore, methods for antibiotic-free plasmid maintenance are in high demand. Here, we present an essential gene-based plasmid selection strategy utilizing the Escherichia coli tryptophan tRNA (trpT) gene. We developed a workflow using a base strain with a trpT deletion and a temperature-sensitive trpT-expressing plasmid to circumvent the need for remaking chromosomal trpT deletions for every transformation. We evaluated the stability of a range of antibiotic gene-free trpT plasmids with different copy numbers and determined that the system is as efficient as, or better than, systems using antibiotics. Furthermore, the system is stable when producing a biochemical at industrially relevant fermentation conditions, and due to the small size of trpT, it allows for plasmid minimization. The approach constitutes a significant contribution toward developing simpler and more effective antibiotic-free bioprocesses and combating the spread of multiresistant infections.

Keywords:  antibiotic-free selection; genetic stability; plasmid minimization; synthetic biology tools; tRNA-based selection; trpT gene

DOI:  https://doi.org/10.1021/acssynbio.5c00622
STAR Protoc. 2025 Dec 22. pii: S2666-1667(25)00686-0. [Epub ahead of print]7(1): 104280

Protocol for marker-free genome editing in Saccharomyces cerevisiae using universal donor templates and multiplexed CRISPR-Cas9.

Darshi Hemani, James H Grissom, Richard J Chi.

  Here, we present a protocol for marker-free genome editing in Saccharomyces cerevisiae by combining PCR-based selectable marker cassettes with CRISPR-Cas9. We describe steps for generating gene deletions using MX6 markers and excising the markers by introducing a reusable guide RNA (gRNA)-Cas9 plasmid and universal repair templates, allowing multiplex removal in a single step. Final verification by PCR yields marker-free strains that can be iteratively edited using the same selectable markers. For complete details on the use and execution of this protocol, please refer to Grissom et al.1.

Keywords:  CRISPR; Cell Biology; Model Organisms

DOI:  https://doi.org/10.1016/j.xpro.2025.104280
Nat Commun. 2025 Dec 27.

Photocontrolled trimethoprim PROTACs targeting the eDHFR protein tag.

Nitika Sharma, Swarbhanu Sarkar, Tongil Ko, Kimberly J Edwards, Jonathan M Pham, Tommy Nguyen, Angela Z Gong, Joseph Flores, Dirk Trauner, Mark A Sellmyer.

Proteolysis targeting chimeric small molecules (PROTACs) offer a strategy for degrading disease-associated proteins or controlling engineered protein tags fused to therapeutic proteins, like chimeric antigen receptors (CARs). New approaches are needed that allow spatiotemporal control of PROTAC activity, restricting degrader activity to targeted cells. Photopharmacology offers a solution by enabling light-mediated spatial control of drug action. Here, we synthesize photocaged and photoswitchable PROTAC molecules and test their regulation of proteins tagged with E. coli dihydrofolate reductase (eDHFR) in tumor and CAR-T cells. Several of the molecules are derived from triazole-linked trimethoprim-PROTACs (TMP-TACtz), that degrade eDHFR fused proteins at picomolar concentrations, show degradation in cells with low cereblon E3 ligase levels, and have little off-target effects. The photocleavable compound, TMP-TAC-PC yields the best light-mediated regulation of CAR T cell cytotoxicity and cytokine secretion. This work introduces photocontrolled, tag-directed degraders for controlling protein expression in tumor cells and CAR T cells.

DOI: https://doi.org/10.1038/s41467-025-67527-5
Nanoscale Horiz. 2025 Dec 23.

DNA-based hydrogels: a promising material for future energy storage applications.

Samanth Kokkiligadda, Surya Kiran Ampasala, Soong Ho Um.

DNA hydrogels have emerged as promising natural biomaterials for next-generation energy storage systems, offering a unique combination of biocompatibility, programmability, tunability, and self-assembly capabilities. Traditionally developed using synthetic DNA strands or DNA origami, efforts are turning toward naturally derived genomic DNA, such as that obtained from salmon sperm, chicken blood, and other biowaste sources, offering a more sustainable and cost-effective route. These hydrogels possess inherent sequence diversity and tunable network structures, making them ideal candidates for enhancing ionic conductivity, mechanical stability, and electrochemical performance in devices like batteries and supercapacitors. This review explores the foundational principles, synthesis strategies, and recent advancements in using DNA hydrogels as components in batteries, supercapacitors, and fuel cells. Compared to traditional materials, DNA hydrogels provide sustainable advantages such as biodegradability, mechanical flexibility, and designable structures that respond to environmental stimuli. While challenges like limited conductivity, stability, and scaling issues remain, ongoing research is addressing these through chemical modifications, hybrid composites, and integration with nanomaterials. Looking ahead, the development of smart, multifunctional DNA hydrogels holds significant potential to transform energy storage technologies and contribute to global sustainability goals. This review highlights key opportunities and calls for interdisciplinary efforts to fully realize the capabilities of DNA hydrogels in future energy systems.

DOI: https://doi.org/10.1039/d5nh00490j
Proc Natl Acad Sci U S A. 2025 Dec 30. 122(52): e2529119122

Getting gene expression "just right".

Paul C Whitford.

DOI: https://doi.org/10.1073/pnas.2529119122
Biophys J. 2025 Dec 23. pii: S0006-3495(25)03507-6. [Epub ahead of print]

Protein force spectroscopy using magnetic tweezers: slow and steady wins the race?

Stefanie D Pritzl, Jan Lipfert.

  Mechanical forces are central to biological function across scales, from whole organisms to individual molecules. At the cellular and subcellular levels, force generation, sensing, and mechanotransduction shape diverse processes including gene expression, morphogenesis, and disease progression. Single-molecule force spectroscopy provides critical insights into these mechanics, with magnetic tweezers (MT) emerging as a versatile tool with unique advantages. MT operate across physiologically relevant forces (∼0.01-100 pN) and enable stable, long-duration, and multiplexed measurements without photodamage, making them ideally suited to investigate proteins under near-native conditions. This review highlights the evolution of MT-based protein mechanics, spanning early cell microrheology to recent single-molecule studies. We focus on key developments and applications, including investigations of cytoskeletal, membrane, and motor proteins, force-sensitive cell adhesion complexes, mechanoresponsive ion channels, and virus-host interactions. Furthermore, we discuss the integration of MT with fluorescence readouts and emerging in vivo applications, underscoring the expanding role of MT in decoding the molecular basis of mechanobiology.

Keywords:  Magnetic tweezers; force-regulation; force-spectroscopy; protein-protein interactions; proteins

DOI:  https://doi.org/10.1016/j.bpj.2025.12.028
Adv Mater. 2025 Dec 26. e14435

Integrating Vibrio natriegens for Photon Manipulation in Living Lighting Devices.

Stephanie Willeit, Maurice Hädrich, Philippe-Quentin Liss, Valeria Rodriguez Espinoza, Nicklas Maximilian Foerster, Bastian Blombach, Rubén D Costa.

  Photon manipulation with bacteria is an emerging field in photonics (lasers, bio-imaging, and cell-sensing) and optoelectronics (bacterial-hybrid light-emitting diodes (BaHLEDs) and photovoltaics). In BaHLEDs, living photon down-conversion filters based on the direct use of bacteria in optically desirable hydrophobic and/or waterless coatings have not been realized yet. Herein, we put forward a simple concept: the engineering of fluorescent Vibrio natriegens (V. natriegens) that enable easy-to-prepare, untreated bacteria-silicone filters for the first red-emitting BaHLEDs. More specifically, we have rationalized genetic and material engineering tools to optimize i) the protein production time with a 1.7-fold enhanced volumetric productivity compared to E. coli with the same spectroscopic quality for the fluorescent protein (FP) DsRed, ii) the straightforward fabrication of highly emissive and stable V. natriegens-silicones, and iii) device reproducibility and performance to reach competitive devices with stabilities ranging from a few days up to weeks depending on device working conditions and architectures. Overall, this work sets in bacterial photon down-conversion using V. natriegens directly as a viable and more straightforward concept toward further advances in living lighting applications.

Keywords:   Vibrio natriegens ; bacteria‐hybrid light‐emitting diodes; engineering living materials; fluorescent proteins; photon management

DOI:  https://doi.org/10.1002/adma.202514435
Nat Commun. 2025 Dec 22.

Computational design of allulose-responsive biosensor toolbox for auto-inducible protein expression and CRISPRi mediated dynamic metabolic regulation.

Qianzhen Dong, Peng Chen, Zhiru Guo, Hongli Wei, Yan Zeng, Jingyu Zhang, Yan Men, Weidong Liu, Yuanxia Sun, Jiangang Yang.

Biosensors based on transcription factors (TFs) have shown extensive applications in synthetic biology. Due to the complex multi-domain structure of effector-TF-DNA, computational design of TFs remains a challenge. Here, we present the successful structure-guided computational design of the access tunnel, ligand binding, allosteric transition process for an allulose-responsive PsiR. It enables a 20-fold increase in sensitivity, reducing the EC50 of PsiR-allulose biosensors (PABs) from 16 mM to 0.8 mM, and delivers a PAB box possessing the detection range from 10 μM to 100 mM. We further validate its broader applicability in enhancing sensitivity of LacI-IPTG biosensor. Based on the developed PABs, we present the inducer-free allulose-mediated auto-inducible protein expression system, and demonstrate an allulose-triggered CRISPR interference circuit for dynamic metabolic regulation. It facilitates a 68% increase in allulose titer and achieves a high yield of 0.43 g/g glucose. This work provides the versatile TF toolbox for developing allulose-triggered regulation circuits in biotechnology application.

DOI: https://doi.org/10.1038/s41467-025-67669-6
Small. 2025 Dec 23. e11147

Responsive Emulsions Enabled by Supramolecular Polymers for Recyclable Biocatalysis.

Jieyao Zhang, Shan Wang, Changzhu Wu.

  Emulsion biocatalysis harnesses liquid-liquid interfaces to facilitate mass transfer for achieving efficient biocatalysis. However, it has been a great challenge in recycling the emulsions in the downstream process. Here, we present a supramolecular approach to design a responsive emulsion for recyclable biocatalysis. Crown ether acrylate monomers are copolymerized with N-isopropylacrylamide to yield amphiphilic supramolecular polymer as emulsifiers, which further enable to form robust emulsions, accommodating both isolated enzymes and whole-cell catalysts and supporting single-step transformations as well as multi-enzymatic cascades, with performance that surpasses conventional biphasic systems. Importantly, leveraging the reversible crown-ether/secondary-ammonium recognition, the introduction of a complementary dibenzylammonium guest triggers supramolecular cross-linking, enabling straightforward recovery and reuse of both emulsifier and catalyst with minimal activity loss, and subsequent mild pH modulation de-complexes the host-guest assembly, resulting in a reversibly switching system between emulsion and two-phase states. Additionally, Escherichia coli overexpressing BAL retains 80% of its initial activity over three consecutive cycles in this responsive emulsion. Therefore, our findings establish supramolecular polymer-stabilized emulsions as a versatile, recyclable platform for biocatalysis, providing a general strategy that bridges supramolecular polymer design with green chemical synthesis and offering strong potential for sustainable industrial chemical synthesis.

Keywords:  biocatalysis; cascade; host‐guest chemistry; responsive emulsion; supramolecular polymer

DOI:  https://doi.org/10.1002/smll.202511147
Proc Natl Acad Sci U S A. 2025 Dec 30. 122(52): e2425774122

Multiplex mapping of protein-protein interaction interfaces.

Jingxuan He, Ling-Nan Zou, Vidhi Pareek, Stephen J Benkovic.

  We describe peptide mapping through Split Antibiotic Resistance Complementation (SpARC-map), a method to identify the probable interface between two interacting proteins. Our method is based on in vivo affinity selection inside a bacterial host and uses high-throughput DNA sequencing to infer probable protein-protein interaction (PPI) interfaces. SpARC-map uses only routine microbiology techniques, with no reliance on specialized instrumentation, dedicated reagents, or reconstituting protein complexes in vitro. SpARC-map can be tuned to detect PPIs over a broad range of affinities, multiplexed to probe multiple PPIs in parallel, and its nonspecific background can be precisely measured, enabling the sensitive detection of weak PPIs. Using SpARC-map, we recover known PPI interfaces in the p21-PCNA, p53-MDM2, and MYC-MAX complexes. We also use SpARC-map to probe the purinosome, the weakly bound complex of six purine biosynthetic enzymes, where no PPI interfaces are known. There, we identify interfaces that satisfy structural requirements for substrate channeling, as well as protein surfaces that participate in multiple distinct interactions, which we validate using site-specific photocrosslinking in live human cells. Finally, we show that SpARC-map results can impose stringent constraints on machine learning-based structure prediction.

Keywords:  protein complex; protein interaction interface; protein–protein interaction; purinosome

DOI:  https://doi.org/10.1073/pnas.2425774122
ACS Synth Biol. 2025 Dec 24.

A Simple and Versatile Cell-Free Expression Method for Producing Secondary Metabolites.

Jaime Lorenzo N Dinglasan, Namil Lee, Nam Ngoc Pham, Meghana Faltane, Marie Lynde, Katherine B Louie, Sangeeta Nath, Jay D Keasling, Hiroshi Otani, Nigel J Mouncey.

  Secondary metabolites are a major source of natural products with industrially relevant bioactivities. Lysate-based cell-free expression (CFE) is an emerging platform for accelerating the discovery and engineering of these natural products. While Escherichia coli cell extracts are widely used for CFE, Streptomyces extracts are likely to offer a more biochemically compatible environment for their expression. However, current Streptomyces-based CFE systems remain underdeveloped, with protocols that are either strain-specific or not readily scalable. To address these limitations and enable broader access to cell-free natural product biosynthesis, we present a generalizable and simple set of reaction conditions that support high-yield protein expression (180-230 μg/mL) in lysates derived from Streptomyces venezuelae NRRL B-65422 and Streptomyces lividans TK24. Like E. coli-based systems, these extracts enable iterative and pathway-level biosynthesis, as demonstrated by the production of the polyketide flaviolin and the cyclic dipeptide albonoursin. Notably, the S. lividans lysate outperforms the E. coli systems by also supporting the expression and catalytic activity of a (∼250 kDa) type I polyketide synthase (T1PKS), producing its corresponding ethyl ketone product, 2-methyl-3-pentanone, without the need for precursor or post-translational modification supplements. To our knowledge, this represents the first demonstration coupling both expression and catalysis of a megasynthase in a Streptomyces-based system, and of a T1PKS in any bacterial extract. By addressing key challenges in the generalizability and scalability of prior Streptomyces CFE, we establish a protocol that enables parallelized evaluation of diverse lysate systems and provides a foundation for high-throughput T1PKS engineering in vitro.

Keywords:  Streptomyces; cell-free expression; natural product synthesis; secondary metabolites; type I polyketide synthase

DOI:  https://doi.org/10.1021/acssynbio.5c00497
Nat Commun. 2025 Dec 22.

Maintenance of cytoplasmic and membrane densities shapes cellular geometry in Escherichia coli.

Griffin Chure, Roshali T de Silva, Richa Sharma, Michael C Lanz, Jonas Cremer.

Microbes precisely control their composition and geometry across diverse growth conditions, yet the mechanisms coordinating these processes remain unclear. Here, we integrate quantitative proteomics, microscopy, and biochemical measurements to reveal a biophysical principle linking these properties in Escherichia coli: cytoplasmic and membrane protein densities maintain a tightly conserved ratio across growth conditions, while the periplasmic density varies. Building on this observation, we develop a mathematical model demonstrating that maintaining this density ratio constrains the surface-to-volume ratio as a nonlinear function of proteome composition, specifically the ribosomal proteome fraction and partitioning between cellular compartments. The model holds under guanosine tetraphosphate perturbations that alter ribosome levels, further demonstrating that cellular geometry is not strictly determined by growth rate. These findings provide a biophysical framework for geometry control, underscoring density maintenance as a key physiological constraint that shapes cellular phenotypes.

DOI: https://doi.org/10.1038/s41467-025-67553-3
Nat Commun. 2025 Dec 23.

Closed-loop optogenetic control of cell biology enables outcome-driven microscopy.

Josiah B Passmore, Alfredo Rates, Jakob Schröder, Menno T P van Laarhoven, Vincent J W Hellebrekers, Henrik G van Hoef, Antonius J M Geurts, Wendy van Straaten, Wilco Nijenhuis, Florian Berger, Carlas S Smith, Ihor Smal, Lukas C Kapitein.

Smart microscopy is transforming biological imaging by integrating real-time analysis with adaptive acquisition to enhance imaging efficiency. Whereas many emerging implementations are event-driven and focus on on-demand data acquisition to reduce phototoxicity, we here present 'outcome-driven' microscopy, a framework combining smart microscopy with optogenetics to control cell biological processes and achieve predefined outcomes. We validate this approach using light-based control of cell migration and nucleocytoplasmic transport, demonstrating robust spatiotemporal control of cellular behaviour in single cells and in cell populations.

DOI: https://doi.org/10.1038/s41467-025-67848-5
Nat Genet. 2025 Dec 23.

Designing synthetic regulatory elements using the generative AI framework DNA-Diffusion.

Lucas Ferreira DaSilva, Simon Senan, Judith F Kribelbauer-Swietek, Zain Munir Patel, Lithin Karmel Louis, Aniketh Janardhan Reddy, Sameer Gabbita, Jonathan D Rosen, Zach Nussbaum, César Miguel Valdez Córdova, Aaron Wenteler, Noah Weber, Tin M Tunjic, Martino Mansoldo, Talha Ahmad Khan, Gue-Ho Hwang, Vincent Gardeux, David T Humphreys, Cameron Smith, Matei Bejan, Peter Bromley, Will Connell, Bart Deplancke, Michael I Love, Emily S Wong, Wouter Meuleman, Luca Pinello.

Systematically designing regulatory elements for precise gene expression control remains a central challenge in genomics and synthetic biology. Here we introduce DNA-Diffusion, a generative artificial intelligence framework that uses machine learning trained on DNA accessibility data from diverse cell lines to design compact regulatory elements with cell-type-specific activity. We show that DNA-Diffusion generates 200-base-pair synthetic elements that recapitulate endogenous transcription factor binding grammar while exhibiting enhanced cell-type specificity. We validated these elements using a 5,850-element STARR-seq library across three cell lines. Moreover, we demonstrated successful endogenous gene modulation using EXTRA-seq, reactivating AXIN2, a leukemia-protective gene, in its native genomic context. Our approach outperforms existing computational methods in balancing functional activity with cell-type specificity while maintaining sequence diversity. This work establishes DNA-Diffusion as a powerful tool for engineering compact, highly specific regulatory elements crucial for advancing gene therapies and understanding gene regulation.

DOI: https://doi.org/10.1038/s41588-025-02441-6
ArXiv. 2025 Dec 19. pii: arXiv:2512.17597v1. [Epub ahead of print]

Design of a minimal, allosteric, and ATPase-like machine using mechanical linkages.

Tosan Omabegho.

ATPases cyclically convert chemical energy in the form of ATP gradients into directed motion inside cells. To function, ATPases rely on allosteric communication between at least two binding sites, an internal signaling mechanism that is not well understood. Here, we model an ATPase-like machine by using a system of mechanical linkages to recreate negative allosteric coupling between two binding sites and generate cycles in which the sites alternate occupancy. The ATPase analog has two mechanical degrees of freedom and two discretized binding sites: one for the ATP, Pi and ADP analogs, and one for an allosteric effector analog. The geometry of the ATPase analog allows stepwise binding reactions at each site to capture the two degrees of freedom in a mutually exclusive way. Consequently, the enzyme interconverts between multiple rigid and partially rigid forms, such that neither site can be fully bound when both sites are occupied. Two mechanisms work together to generate an enzymatic cycle: one, in which the tighter-binding ATP analog can bind and displace the effector from the enzyme; and a second, in which flexibility introduced by splitting the ATP analog into two pieces (catalysis) allows the effector to rebind and displace the products (ADP analog). We show that cleavage (forward catalysis) and ligation (reverse catalysis) alter the rigidity of the enzyme complex equivalently to binding and dissociation, respectively, but must do so more slowly for effective cycling to take place. Simple designs for synthetic systems that mimic ATPase monomers can be derived from this work.
J Am Chem Soc. 2025 Dec 22.

Genetic Code Expansion in Probiotics Enables the Secretion of Covalent Protein Drugs in Mice.

Kaiyi Wang, Yumeng Wu, Jiamin Shang, Lingxin Bu, Da Hu, Honglin He, Ruixuan Wang, Junyi Mao, Yao Yan, Jun Guan, Rui Wang, Huixin Luo.

Oral delivery of therapeutic proteins remains a formidable challenge. Although engineered microbes have emerged as promising platforms for localized drug synthesis in the gut, their functional capacity has been restricted to the 20 canonical amino acids, limiting the chemical diversity of biologic payloads. Here, we demonstrate that integrating genetic code expansion (GCE) into bacterial therapy overcomes this fundamental constraint. We engineered the probiotic Escherichia coli Nissle 1917 (EcN) to incorporate the noncanonical amino acid fluorosulfate-l-tyrosine (FSY), enabling in situ secretion of a site-specifically modified covalent anti-IL-23 nanobody exhibiting picomolar binding potency (5.9 pM). Oral administration of this engineered EcN strain, followed by FSY supplementation, significantly ameliorated colitis in a murine model. This approach thereby establishes a versatile and generalizable platform that substantially expands the functional scope and therapeutic potential of live biotherapeutics.

DOI: https://doi.org/10.1021/jacs.5c17978
ACS Synth Biol. 2025 Dec 22.

High-Throughput Detection of Cyanobacterial Form I Rubisco Assembly.

Jackson W Wysocki, ByungUk Lee, Tina Wang.

  Rubisco catalyzes the CO2 fixation step in the dark reactions of photosynthesis. Transgenic expression of better-performing Rubisco orthologs in plants or discovery of improved mutants of Rubisco via protein engineering could theoretically accelerate plant growth and improve crop yields. However, efforts to heterologously express or engineer Rubisco are frequently stymied by the chaperone-dependent folding and assembly of the Rubisco holoenzyme, a process that can be disrupted by changes to Rubisco's sequence. Elucidation of the effects that alterations to Rubisco's sequence impose upon its biogenesis is hampered by reliance upon low-throughput methods for verification of Rubisco assembly. Here, we report the engineering of a genetically encoded biosensor to sense the assembly of Form I Rubiscos in E. coli. We show that the biosensor can detect the RbcS-dependent assembly of cyanobacterial Rubisco orthologs, the formation of chaperone-stabilized RbcL oligomeric assembly intermediates, and differences in assembly caused by mutations to the RbcL sequence. Additionally, we perform a large-scale examination of the relative assembly levels of a ∼7500-member Halothiobacillus neapolitanus RbcL mutant library by adapting the biosensor for use with phage-assisted noncontinuous selection. Our experiment predicts that the majority (>90%) of examined RbcL mutations exert a negative effect on assembly, lending support to the hypothesis that Rubisco biogenesis constrains both its natural evolution and improvement by protein engineering.

Keywords:  Chaperone−dependent assembly; Genetically encoded biosensor; Protein engineering constraints; RbcL/RbcS assembly; Rubisco biogenesis

DOI:  https://doi.org/10.1021/acssynbio.5c00591