bims-enlima 2025-03-09 papers

bims-enlima

Biomed News

on Engineered living materials

Issue of 2025–03–09
forty-five papers selected by
Rahul Kumar, Tallinna Tehnikaülikool

Designer mammalian living materials through genetic engineering.
Engineering Plasmids with Synthetic Origins of Replication.
3-D Printable Living Hydrogels as Portable Bio-energy Devices.
Controlled macroscopic shape evolution of self-growing polymeric materials.
3D Printed Materials with Nanovoxelated Elastic Moduli.
ATPS-enabled single-step printing of chemically and mechanically on-demand tunable perfusable channels in ejectable constructs.
Measurement and Comparison of Hyaluronic Acid Hydrogel Mechanics Across Length Scales.
Heterovalent Click Reactions on DNA Origami.
Mechano-diffusion of particles in stretchable hydrogels.
Construction of complex bacteriogenic protocells from living material assembly.
Thermally Driven Dynamic Behaviors in Polymeric Vesicles.
Microbial Production of High-Performance Fibers from Muscle Protein Titin.
Diels-Alder-based IPN hydrogels with tunable mechanical and protein release properties for tissue engineering.
Enhanced Cell Proliferation and Maturation Using Carboxylated Bacterial Nanocellulose Scaffolds for 3D Cell Culture.
Cell-Free Expression of Soluble Leafhopper Proteins from Brochosomes.
Programming Nutrient Detection with Modular Regulators for Dynamic Control of Microbial Biosynthesis.
Genome duplication in a long-term multicellularity evolution experiment.
Controlling the Dynamic Behavior of Microposts in Solution via Diffusion-Convection.
Post-assembly Plasmid Amplification for Increased Transformation Yields in E. coli and S. cerevisiae.
Nucleic Acid Framework-Enabled Spatial Organization for Biological Applications.
Biomimetic Chemocatalytic Cascades-A New Strategy for Molecular Design of Degradable Polymer Systems.
Sunlight-Driven Photomobile Polymer Materials Containing Push-Pull Azobenzene Moieties.
Protocol to modulate SUMOylation of a specific protein in budding yeast using chemical genetic approaches.
Liquid Printing in Nanochitin Suspensions: Interfacial Nanoparticle Assembly Toward Volumetric Elements, Organic Electronics and Core-Shell Filaments.
Adhesion study at the interface of a PDMS-elastomer and borosilicate glass-slide: effect of modulus and thickness of the elastomer.
High-resolution bioprinting of complex bio-structures via engineering of the photopatterning approaches and adaptive segmentation.
AI-enabled manufacturing process discovery.
Hybrid Ultrathin Gold Nanowire Gels: Formation and Mechanical Properties.
In Situ-Gelling Antimicrobial Poly(oligoethylene glycol methacrylate)-Based Hydrogels Integrating Bound Quaternary Ammonia Compounds and Antibiotic Functionalities for Effective Infected Wound Healing.
Controlled Enzyme Cargo Loading in Engineered Bacterial Microcompartment Shells.
Micropores can enhance the intrinsic fracture energy of hydrogels.
A microfluidic platform for extraction and analysis of bacterial genomic DNA.
Cell size regulation in bacteria: a tale of old regulators with new mechanisms.
Computation-aided designs enable developing auxotrophic metabolic sensors for wide-range glyoxylate and glycolate detection.
Viscoelastic mechanics of living cells.
One-step Cre-loxP organism creation by TAx9.
Capturing CDKs in Action: Live-Cell Biosensors Pioneer the New Frontiers in Cell Cycle Research.
Bacteria-Mediated Intracellular Radical Polymerizations.
Facile Synthesis of Silicone Oil-Based Ferrofluid: Toward Smart Materials and Soft Robots.
Overcoming ice: cutting-edge materials and advanced strategies for effective cryopreservation of biosample.
Regulation of gene expression through protein-metabolite interactions.
Synthetic Biology-Based Strategy for Nanomedicine Design.
From Unprintable Peptidic Gel to Unstoppable: Transforming Diphenylalanine Peptide (Fmoc-FF) Nanowires and Cellulose Nanofibrils into a High-Performance Biobased Gel for 3D Printing.
Presenting Antimicrobial Peptides on Poly(ethylene glycol): Star-Shaped vs Comb-Like Architectures.
Site-Specific Polymer-Protein-Polymer Conjugates for the Preparation of Dual Responsive Multilayer Nanoparticles.

Bioact Mater. 2025 Jun;48 135-148

Designer mammalian living materials through genetic engineering.

Mariana Gameiro, José Almeida-Pinto, Beatriz S Moura, João F Mano, Vítor M Gaspar.

  Emerging genome editing and synthetic biology toolboxes can accurately program mammalian cells behavior from the inside-out. Such engineered living units can be perceived as key building blocks for bioengineering mammalian cell-dense materials, with promising features to be used as living therapeutics for tissue engineering or disease modeling applications. Aiming to reach full control over the code that governs cell behavior, inside-out engineering approaches have potential to fully unlock user-defined living materials encoded with tailored cellular functionalities and spatial arrangements. Dwelling on this, herein, we discuss the most recent advances and opportunities unlocked by genetic engineering strategies, and on their use for the assembly of next-generation cell-rich or cell-based materials, with an unprecedent control over cellular arrangements and customizable therapeutic capabilities. We envision that the continuous synergy between inside-out and outside-in cell engineering approaches will potentiate the future development of increasingly sophisticated cell assemblies that may operate with augmented biofunctionalities.

Keywords:  Genetic engineering; Living materials; Mammalian cells; Synthetic biology; Tissue engineering

DOI:  https://doi.org/10.1016/j.bioactmat.2025.02.007
bioRxiv. 2025 Feb 21. pii: 2025.02.21.639468. [Epub ahead of print]

Engineering Plasmids with Synthetic Origins of Replication.

Baiyang Liu, Xiao Peng, Matthew R Bennett, Matthew R Lakin, James Chappell.

Plasmids remain by far the most common medium for delivering engineered DNA to microorganisms. However, the reliance on natural plasmid replication mechanisms limits their tunability, compatibility, and modularity. Here we refactor the natural pMB1 origin and create plasmids with customizable copy numbers by tuning refactored components. We then create compatible origins that use synthetic RNA regulators to implement independent copy control. We further demonstrate that the synthetic origin of replication (SynORI) can be engineered modularly to respond to various signals, allowing for multiplexed copy-based reporting of environmental signals. Lastly, a library of 6 orthogonal SynORI plasmids is created and co-maintained in E. coli for a week. This work establishes the feasibility of creating plasmids with SynORI that can serve as a new biotechnology for synthetic biology.

DOI: https://doi.org/10.1101/2025.02.21.639468
Adv Mater. 2025 Mar 05. e2419249

3-D Printable Living Hydrogels as Portable Bio-energy Devices.

Xinyu Wang, Fei Han, Zhe Xiao, Xiaomeng Zhou, Xingwu Liu, Yue Chen, Ke Li, Yuanheng Li, Qianhengyuan Yu, Hang Zhao, Minshen Zhu, Renheng Wang, Zhiyuan Liu, Chao Zhong.

  Harnessing engineered living materials for energy application represents a promising avenue to sustainable energy conversion and storage, with bio-batteries emerging as a pivotal direction for sustainable power supply. Whereas, the realization of miniaturized and portable bio-battery orchestrating off-the-shelf devices remains a significant challenge. Here, this work reports the development of a miniaturized and portable bio-battery using living hydrogels containing conductive biofilms encapsulated in an alginate matrix for nerve stimulation. These hydrogels, which can be 3-D printed into customized geometries, retained biologically active characteristics, including electroactivity that facilitates electron generation and the reduction of graphene oxide. By fabricating the living hydrogel into a standard 2032 battery shell with a diameter of 20 mm, this work successfully creates a miniaturized and portable bio-battery with self-charging performance. The device demonstrates remarkable electrochemical performance with a coulombic efficiency of 99.5% and maintains high cell viability exceeding 90% after operation. Notably, the electricity generated by the bio-battery can be harnessed for nerve stimulation to enable precise control over bioelectrical stimulation and physiological blood pressure signals. This study paves the way for the development of novel, compact, and portable bio-energy devices with immense potential for future advancements in sustainable energy technologies.

Keywords:  bio‐energy device; engineered living energy materials; materials synthetic biology; miniaturized and portable bio‐battery; nerve stimulation

DOI:  https://doi.org/10.1002/adma.202419249
Nat Commun. 2025 Mar 03. 16(1): 2131

Controlled macroscopic shape evolution of self-growing polymeric materials.

Xinhong Xiong, Xiaozhuang Zhou, Haohui Zhang, Michael Aizenberg, Yuxing Yao, Yuhang Hu, Joanna Aizenbergd, Jiaxi Cui.

Living organisms absorb external nutrients to grow, changing their macroscopic shapes to meet various challenges through mass transport and integration. While several strategies have been developed to create dynamic polymers that allow for mainchain remodelings to mimic the growing ability of living organisms, most are limited to simple homogeneous growth without complex control of global geometric transformation during growth. Herein, we report an approach to design controlled, growth-induced shape transformation in synthetic materials, in which significant mass transport within the materials is induced by spatially controlled polymerization leading to reshaping the materials. This method is demonstrated using silicone systems made through anionic ring-opening polymerization (anionic ROP) of octamethylcyclotetrasiloxane (D4) with a strong base as the catalyst. We show that a flat square sample can be transformed into a sphere through growth without the need for remolding and preprogramming. By varying the composition of the monomer mixture provided to the samples, and the modes of triggering and shutting down polymerization, we achieve exquisite control over growing polymeric objects into various sizes and shapes, modulating their mechanical properties, self-healing ability, and availability of active sites for further growth from a desired location. We envision this strategy opening an innovative direction in preparing soft materials with specific shapes or surface morphologies.

DOI: https://doi.org/10.1038/s41467-025-57030-2
Adv Mater. 2025 Mar 06. e2416262

3D Printed Materials with Nanovoxelated Elastic Moduli.

Peter L H Newman, Mohammad Mirkhalaf, Steven C Gauci, Iman Roohani, Maté Biro, Christopher Barner-Kowollik, Hala Zreiqat.

  Fabrication methods that synthesize materials with higher precision and complexity at ever smaller scales are rapidly developing. Despite such advances, generating complex 3D materials with controlled mechanical properties at the nanoscale remains challenging. Exerting precise control over mechanical properties at the nanoscale would enable material strengths near theoretical maxima, and the replication of natural structures with hitherto unattainable strength-to-weight ratios. Here, a method for fabricating materials with nanovoxelated elastic moduli by employing a volume-conserving photoresist composed of a copolymer hydrogel, along with OpenScribe, an open-source software that enables the precise programming of material mechanics, is presented. Combining these, a material composed of periodic unit cells featuring heteromechanically tessellated soft-stiff structures, achieving a mechanical transition over an order-of-magnitude change in elastic modulus within 770 nm, a 130-fold improvement on previous reports, is demonstrated. This work critically advances material design and opens new avenues for fabricating materials with specifically tailored properties and functionalities through unparalleled control over nanoscale mechanics.

Keywords:  3d printing; architectured materials; elastic modulus; metamaterials; nanoscale materials

DOI:  https://doi.org/10.1002/adma.202416262
Biofabrication. 2025 Mar 05.

ATPS-enabled single-step printing of chemically and mechanically on-demand tunable perfusable channels in ejectable constructs.

Malin Becker, Francisca Luísa Fernandes Gomes, Isa Robin Porsul, Jeroen Leijten.

  3D bioprinting approaches offer highly versatile solutions to replicate living tissue and organ structures. While current bioprinting approaches can generate desired shapes and spatially determined patterns, the material selection for embedded bioprinting has remained limited, as it has relied on the use of viscous, shear-thinning, or liquid-like solid materials to create shape controlled constructs, which could then be modified downstream via multi-step processes. We here explore aqueous two-phase system stabilized 3D bioprinting of low viscous materials in combination with supramolecular complexation to fabricate intricate, perfusable engineered constructs that are both mechanically and chemically tunable in a single-step manner. To this end, we introduce Dex-TAB as a highly versatile backbone, that allows for mechanical and chemical tuning during as well as after printing. Showcasing the printability as well as spatial chemical modification and mechanical tunability of this material, ejectability, and local/gradual or bulk functionalized interconnected tube shaped constructs were generated. Subsequently, we demonstrated that these functionalized channels could be printed directly into a syringe containing crosslinkable polymer solution, which upon ejection forms pre-patterned perfusable constructs. In short, we report that ATPS enabled low viscous 3D bioprinting can produce highly functional and even potentially minimally invasive injectable yet functionalized and perfusable constructs, which offers opportunities to advance various biofabrication applications.

Keywords:  Embedded bioprinting; biofabrication; biofunctionalization; tissue engineering; vascularization

DOI:  https://doi.org/10.1088/1758-5090/adbcdc
J Biomed Mater Res A. 2025 Mar;113(3): e37889

Measurement and Comparison of Hyaluronic Acid Hydrogel Mechanics Across Length Scales.

Aina Solsona-Pujol, Nikolas Di Caprio, Hannah M Zlotnick, Matthew D Davidson, Morgan B Riffe, Jason A Burdick.

  Hydrogels are an important class of biomaterials that are being developed for use in medicine, such as in drug delivery and tissue engineering applications. To improve properties (e.g., injectability, nutrient transport, cell invasion), hydrogels are often processed as hydrogel microparticles (microgels) that can be used as suspensions or jammed into granular hydrogels. The mechanical properties of microgels are important across length scales, from macroscale bulk properties of granular assemblies to microscale interactions with cells; however, microgel mechanics are rarely reported due to challenges in their measurement. To address this, we report here a cost-effective, easy-to-use do-it-yourself (DIY) active feedback micropipette aspiration device to quantify the mechanics of individual microgels. Using norbornene-modified hyaluronic acid (NorHA) synthesized via an environmentally friendly, aqueous reaction as an exemplary hydrogel, we compare hydrogel mechanics across scales at various macromer concentrations. Hydrogels tested via uniaxial compression exhibit similar moduli values, trends of increasing modulus with increasing macromer concentration, and mechanical stability over time to the same formulations processed as microgels via batch emulsions (~170 μm) and tested via micropipette aspiration. Moduli range from ~50 to ~100 kPa as the NorHA macromer concentration increases from 3 wt% to 5 wt%. These findings are validated by testing with spherical nanoindentation, with similar moduli measured. Collectively, this work provides an accessible device that allows for the rapid testing of microgel mechanical properties, while also improving our understanding of hydrogel mechanics across scales for use in the design of microgels for biomedical applications.

Keywords:  hydrogel; microgel; microgel mechanics; micropipette aspiration; nanoindentation

DOI:  https://doi.org/10.1002/jbm.a.37889
Bioconjug Chem. 2025 Mar 05.

Heterovalent Click Reactions on DNA Origami.

Grant A Knappe, Jeffrey Gorman, Andrew N Bigley, Steven P Harvey, Mark Bathe.

Nucleic acid nanoparticles (NANPs) fabricated by using the DNA origami method have broad utility in materials science and bioengineering. Their site-specific, heterovalent functionalization with secondary molecules such as proteins or fluorophores is a unique feature of this technology that drives its utility. Currently, however, there are few chemistries that enable fast, efficient covalent functionalization of NANPs with a broad conjugate scope and heterovalency. To address this need, we introduce synthetic methods to access inverse electron-demand Diels-Alder chemistry on NANPs. We demonstrate a broad conjugate scope, characterize application-relevant kinetics, and integrate this new chemistry with strain-promoted azide-alkyne cycloaddition chemistry to enable heterovalent click reactions on NANPs. We applied these chemistries to formulate a prototypical chemical countermeasure against chemical nerve agents. We envision this additional chemistry finding broad utility in the synthetic toolkit accessible to the nucleic acid nanotechnology community.

DOI: https://doi.org/10.1021/acs.bioconjchem.4c00552
Soft Matter. 2025 Mar 03.

Mechano-diffusion of particles in stretchable hydrogels.

Chuwei Ye, Congjie Wei, Jiabin Liu, Tsz Hung Wong, Xinyue Liu, Ziyou Song, Chenglin Wu, Zhaojian Li, Shaoting Lin.

Precise control over particle diffusion is promising for diverse modern technologies. Traditionally, particle diffusion is governed by the inherent properties of a liquid medium, limiting versatility and controllability. Here, we report a mechano-diffusion mechanism that harnesses mechanical deformation to control particle diffusion in stretchable hydrogels with a significantly enlarged tuning ratio and a highly expanded tuning freedom. The working principle is to leverage the mechanical deformation of stretchable hydrogels for modulating the polymer network's geometric transformation and the polymer chain's energy modulation, which synergistically tunes the energy barrier for particle diffusion. Using a model particle-hydrogel material system and a customized mechano-diffusion characterization platform, we demonstrate that tension loads can enhance the diffusivity of gold nanoparticles up to 22 times, far exceeding that in traditional liquid medium and by external fields. Additionally, we show particle diffusion in hydrogels can be manipulated spatiotemporally by controlling the hydrogels' stress state and loading rate. To further push the limit of the mechano-diffusion, we use experiment, theory, and simulation to explore particle diffusion in biaxially stretched hydrogels, simultaneously expanding the mesh size and reducing the energy barrier. The enlarged tuning ratio and expanded tuning freedom enable a model-guided drug delivery system for pressure-controlled release of drug molecules. Understanding this spatiotemporal mechano-diffusion mechanism will provide insights pertinent to a broad range of biological and synthetic soft materials.

DOI: https://doi.org/10.1039/d4sm01522c
Nat Protoc. 2025 Mar 05.

Construction of complex bacteriogenic protocells from living material assembly.

Can Xu, Mei Li, Nicolas Martin, Stephen Mann.

Protocell research offers diverse opportunities to understand cellular processes and the foundations of life and holds attractive potential applications across various fields. However, it is still a formidable task to construct a true-to-life synthetic cell with high organizational and functional complexity. Here we present a protocol for constructing bacteriogenic protocells by employing prokaryotes as on-site repositories of compositional, functional and structural building blocks to address this challenge. This approach is based on the capture and processing of two spatially segregated bacterial colonies within individual coacervate microdroplets to produce membrane-bounded, molecularly crowded, compositionally, structurally and functionally complex synthetic cells. The bacteriogenic protocells inherit sufficient biological components from their bacterial building units to exhibit highly integrated life-like properties, including biocatalysis, glycolysis and gene expression. The protocells can be endogenously remodeled to acquire diverse proto-organelles including a spatially partitioned nucleus-like DNA/histone-based condensate to store genetic material, membrane-bounded water vacuoles to adjust cellular osmotic pressure, a three-dimensional network of F-actin proto-cytoskeleton to support structural stability and proto-mitochondria to generate endogenous ATP as source of energy. The protocells ultimately develop a nonspherical morphology due to the continuous biogeneration of metabolic products by implanted living bacteria cells. This protocol provides a novel living material assembly strategy for the construction of functional protoliving microdevices and offers opportunities for potential applications in engineered synthetic biology and biomedicine. The protocol takes ~27 d to complete and requires expertise in microbiology, phase separation, biochemistry and molecular biology related techniques.

DOI: https://doi.org/10.1038/s41596-025-01148-6
Small. 2025 Mar 05. e2411220

Thermally Driven Dynamic Behaviors in Polymeric Vesicles.

Matthew E Allen, Yeyang Sun, Chi Long Chan, Miguel Paez-Perez, Oscar Ces, Yuval Elani, Claudia Contini.

  Stimuli-responsive polymeric vesicles offer a versatile platform for mimicking dynamic cell-like behaviors for synthetic cell applications. In this study, thermally responsive polymeric droplets derived from poly(ethylene oxide)-poly(butylene oxide) (PEO-PBO) polymersomes, aiming to create synthetic cell models that mimic key biological functions are developed. Upon heating, the nanoscale vesicles undergo fusion, transforming into sponge-like microscale droplets enriched with membrane features. By modulating the temperature, these droplets display dynamic properties such as contractility, temperature-induced fusion, and cargo trapping, including small molecules and bacteria, thereby demonstrating their ability to dynamically interface with biological entities. The findings demonstrate the potential of our sponge-like droplets in synthetic cell applications, contributing to the understanding of PEO-PBO polymersomes' unique characteristics, expanding the capabilities of synthetic cell structures, and representing an exciting possibility for advancing soft matter engineering to cell-like behaviors.

Keywords:  biomimicry; polymersomes; self‐assembly; synthetic cells; thermally responsive

DOI:  https://doi.org/10.1002/smll.202411220
Methods Mol Biol. 2025 ;2902 161-172

Microbial Production of High-Performance Fibers from Muscle Protein Titin.

Cameron J Sargent, Christopher H Bowen, Fuzhong Zhang.

  Nature has produced a variety of proteinaceous materials, each with a set of mechanical properties tuned by evolution to adapt to particular environments. While these advantageous properties have also made many of these materials well-suited to various human needs, few protein materials can be harvested from their natural hosts at scale. To meet the demand for these materials using scalable biomanufacturing processes, our lab has developed tools and a biopolymerization platform for the microbial synthesis and processing of nature-derived, high-molecular-weight protein polymers. In this chapter, we describe the application of this platform for polymerizing a segment of the muscle protein titin and processing the resulting polymer into high-performance, muscle-mimetic fibers with a unique combination of desirable mechanical properties.

Keywords:  In vivo polymerization; Protein materials; Synthetic biology

DOI:  https://doi.org/10.1007/978-1-0716-4402-7_10
Int J Biol Macromol. 2025 Mar 04. pii: S0141-8130(25)02330-X. [Epub ahead of print] 141779

Diels-Alder-based IPN hydrogels with tunable mechanical and protein release properties for tissue engineering.

Farouk Segujja, Gökhan Duruksu, Elif Beyza Eren, Aygun İsayeva, Yusufhan Yazır, Ahmet Erdem.

  Advancing hydrogel technology with tunable mechanical strength and sustained release is critical for therapeutic applications in drug delivery and tissue engineering. Conventional single polymer networks, including semi-interpenetrating polymer network (SIPN) hydrogels, often lack mechanical robustness and controlled release needed for therapeutic use. In this study, we fabricated a biocompatible interpenetrating polymer network (IPN) hydrogel with improved properties for controlled protein release. We employed a facile one-pot synthesis approach that integrated aqueous Diels-Alder (DA) 'click' chemistry with photopolymerization methods to crosslink gelatin methacryloyl (GelMA) within a polymeric framework of poly(ethylene) glycol bismaleimide (PEGMI) and multi-furan-modified polyethylene glycol (PEGFU). Spectroscopy (FTIR and 1H NMR) confirmed the chemical composition of the hydrogels. The effect of varying polymer ratios on hydrogel properties was assessed to optimize protein release and mechanical behavior. Fully crosslinked IPN hydrogels exhibited enhanced energy dissipation and compressive moduli 2.5- to 3.5-fold relative to SIPN hydrogels across various polymer ratios. Release kinetics followed the Korsmeyer-Peppas mathematical model, indicating sustained release. IPN hydrogels demonstrated good water absorption, moderate degradation, and favorable biocompatibility with 3 T3 fibroblast cells. Overall, these findings highlight the potential of IPN hydrogels as a promising drug delivery platform for advancing regenerative therapies and targeted treatment strategies.

Keywords:  Diels-Alder reaction; Sustained drug release; Tissue engineering

DOI:  https://doi.org/10.1016/j.ijbiomac.2025.141779
ACS Appl Mater Interfaces. 2025 Mar 05.

Enhanced Cell Proliferation and Maturation Using Carboxylated Bacterial Nanocellulose Scaffolds for 3D Cell Culture.

Elizabeth Mavil-Guerrero, José Manuel Romo-Herrera, Priscila Quiñonez-Angulo, Francisco J Flores-Ruiz, Edén Morales-Narváez, J Félix Armando Soltero, Josué D Mota-Morales, Karla Juarez-Moreno.

  Developing scaffolds for three-dimensional (3D) cell culture and tissue regeneration with biopolymers requires the creation of an optimal nanobiointerface. This interface must possess suitable surface chemistry, biomechanical properties, and fibrillar morphology across nano- to microscale levels to support cell attachment and growth, enabling a biomimetic arrangement. In this study, we developed a hydrogel scaffold made from bacterial nanocellulose (BNC) functionalized with carboxylic acid groups (BNC-COOH) through a reactive deep eutectic solvent (DES), offering a sustainable approach. The surface properties and fibrillar structure of BNC-COOH facilitated the formation of hydrogels with significantly enhanced water uptake (1.4-fold) and adhesion force (2.3-fold) compared to BNC. These hydrogels also demonstrated tissue-like rheological properties in both water with G' exceeding G″, suggesting predominantly elastic (solid-like) characteristics and viscosities in the range of 8-15 Pa·s. The BNC-COOH hydrogel scaffold demonstrated excellent biocompatibility, supporting significant cell growth and anchorage for the 3D growth of mammalian cells and enhancing preadipocyte growth by up to 7.3 times. Furthermore, the BNC-COOH hydrogel facilitates the maturation of 3T3-L1 preadipocytes into mature adipocytes, inducing typical morphology changes, such as decreased filopodia extensions, rounded cell shape, and lipid droplet accumulation without any additional chemical induction stimulus. Therefore, we demonstrated that a reactive DES composed of oxalic acid and choline chloride represents a mild reaction medium and a suitable approach for designing biocompatible 3D hydrogel scaffolds with improved physicochemical properties and biological activities for 3D cell culture.

Keywords:  3D scaffold; 3T3-L1 cells; DES; carboxylated bacterial nanocellulose; cell culture; cell maturation; nanotoxicology

DOI:  https://doi.org/10.1021/acsami.4c22475
ACS Synth Biol. 2025 Mar 07.

Cell-Free Expression of Soluble Leafhopper Proteins from Brochosomes.

Caleb G Lay, Gabriel R Burks, Zheng Li, Jeffrey E Barrick, Charles M Schroeder, Ashty S Karim, Michael C Jewett.

  Brochosomes are proteinaceous nanostructures produced by leafhopper insects with superhydrophobic and antireflective properties. Unfortunately, the production and study of brochosome-based materials has been limited by poor understanding of their major constituent subunit proteins, known as brochosomins, as well as their sensitivity to redox conditions due to essential disulfide bonds. Here, we used cell-free gene expression (CFE) to achieve recombinant production and analysis of brochosomin proteins. Through the optimization of redox environment, reaction temperature, and disulfide bond isomerase concentration, we achieved soluble brochosomin yields of up to 341 ± 30 μg/mL. Analysis using dynamic light scattering and transmission electron microscopy revealed distinct aggregation patterns among cell-free mixtures with different expressed brochosomins. We anticipate that the CFE methods developed here will accelerate the ability to change the geometries and properties of natural and modified brochosomes, as well as facilitate the expression and structural analysis of other poorly understood protein complexes.

Keywords:  Cell-free protein synthesis; brochosomes; disulfide bonds; protein folding; protein nanostructures; self-assembly

DOI:  https://doi.org/10.1021/acssynbio.4c00773
ACS Synth Biol. 2025 Mar 04.

Programming Nutrient Detection with Modular Regulators for Dynamic Control of Microbial Biosynthesis.

Nhu Nguyen, Vincenzo Kennedy, Jung Yeon Lee, Noel Y Chan, Clement T Y Chan.

  Dynamic control of biosynthetic pathways improves the bioproduction efficiency. One common approach is to use genetic sensors that control pathway expression in response to a nutrient molecule in the target feedstock. However, programming the cellular response requires the engineering of numerous genetic parts, which poses a significant barrier to explore the use of different nutrients as cellular signals. Here we created a dynamic control platform based on a set of modular transcriptional regulators; these regulators control the same promoter for driving gene expression, but each of them responds to a unique signal. We demonstrated that by replacing only the regulator, a different nutrient molecule can then be used for induction of the same genetic circuit. To show host versatility, we implemented this platform in both Escherichia coli and Pseudomonas putida. This platform was then used to program the induction of ethanol production by three nutrients, fructose, cellobiose, and galactose, of which each molecule can be present in a different set of crops. These results suggest that our platform facilitates the use of different agricultural products for the dynamic control of biosynthesis.

Keywords:  biosynthesis; dynamic control of metabolic pathway; genetic sensor; microbial engineering

DOI:  https://doi.org/10.1021/acssynbio.4c00720
Nature. 2025 Mar 05.

Genome duplication in a long-term multicellularity evolution experiment.

Kai Tong, Sayantan Datta, Vivian Cheng, Daniella J Haas, Saranya Gourisetti, Harley L Yopp, Thomas C Day, Dung T Lac, Ahmad S Khalil, Peter L Conlin, G Ozan Bozdag, William C Ratcliff.

Whole-genome duplication (WGD) is widespread across eukaryotes and can promote adaptive evolution1-4. However, given the instability of newly formed polyploid genomes5-7, understanding how WGDs arise in a population, persist, and underpin adaptations remains a challenge. Here, using our ongoing Multicellularity Long Term Evolution Experiment (MuLTEE)8, we show that diploid snowflake yeast (Saccharomyces cerevisiae) under selection for larger multicellular size rapidly evolve to be tetraploid. From their origin within the first 50 days of the experiment, tetraploids persisted for the next 950 days (nearly 5,000 generations, the current leading edge of our experiment) in 10 replicate populations, despite being genomically unstable. Using synthetic reconstruction, biophysical modelling and counter-selection, we found that tetraploidy evolved because it confers immediate fitness benefits under this selection, by producing larger, longer cells that yield larger clusters. The same selective benefit also maintained tetraploidy over long evolutionary timescales, inhibiting the reversion to diploidy that is typically seen in laboratory evolution experiments. Once established, tetraploidy facilitated novel genetic routes for adaptation, having a key role in the evolution of macroscopic multicellular size via the origin of evolutionarily conserved aneuploidy. These results provide unique empirical insights into the evolutionary dynamics and impacts of WGD, showing how it can initially arise due to its immediate adaptive benefits, be maintained by selection and fuel long-term innovations by creating additional dimensions of heritable genetic variation.

DOI: https://doi.org/10.1038/s41586-025-08689-6
Langmuir. 2025 Mar 04.

Controlling the Dynamic Behavior of Microposts in Solution via Diffusion-Convection.

Moslem Moradi, Oleg E Shklyaev, Anna C Balazs.

Solutal buoyancy forces in solution arise from density gradients, which occur when the reactants and products of a chemical reaction occupy different volumes in the fluid. These forces drive fluids to spontaneously perform self-directed mechanical work such as shaping and organizing flexible objects in fluid-filled microchambers. Here, we use theory and simulation to show that chemical reactions are not necessary to generate useful solutal buoyancy forces; it is sufficient to just add reactants to aqueous solutions that have a different mass-to-volume ratio than water to drive such spontaneous mechanical action. To demonstrate this behavior, we model arrays of tethered, nonreactive posts in a fluid-filled chamber. Relatively dense chemicals released from the chamber's side walls diffuse into the solution and generate buoyancy-driven flows, which spontaneously trigger the posts to undergo collective dynamics. The posts' dynamics can be controllably programmed by staging the sequence of chemical release from the different walls. With chemically active posts within the array, turning on and off the influx of chemicals from the side walls leads to propagating waves that drive the posts to exhibit biomimetic coordinated motion. The introduction of cascade reactions dynamically shifts the direction of wave propagation. Our findings show how diffusion-convection and diffusion-reaction-convection processes can be used to regulate nonequilibrium spatiotemporal behavior in fluidic systems. This level of control is vital for creating portable microfluidic devices that operate without external power sources and thus function in remote or resource-poor locations.

DOI: https://doi.org/10.1021/acs.langmuir.4c04567
Chem Bio Eng. 2025 Feb 27. 2(2): 87-96

Post-assembly Plasmid Amplification for Increased Transformation Yields in E. coli and S. cerevisiae.

Thomas Fryer, Darian S Wolff, Max D Overath, Elena Schäfer, Andreas H Laustsen, Timothy P Jenkins, Carsten Andersen.

Many biological disciplines rely upon the transformation of host cells with heterologous DNA to edit, engineer, or examine biological phenotypes. Transformation of model cell strains (Escherichia coli) under model conditions (electroporation of circular supercoiled plasmid DNA; typically pUC19) can achieve >1010 transformants/μg DNA. Yet outside of these conditions, e.g., work with relaxed plasmid DNA from in vitro assembly reactions (cloned DNA) or nonmodel organisms, the efficiency of transformation can drop by multiple orders of magnitude. Overcoming these inefficiencies requires cost- and time-intensive processes, such as generating large quantities of appropriately formatted input DNA or transforming many aliquots of cells in parallel. We sought to simplify the generation of large quantities of appropriately formatted input cloned DNA by using rolling circle amplification (RCA) and treatment with specific endonucleases to generate an efficiently transformable linear DNA product for in vivo circularization in host cells. We achieved an over 6500-fold increase in the yield of input DNA, and demonstrate that the use of a nicking endonuclease to generate homologous single-stranded ends increases the efficiency of E. coli chemical transformation compared to both linear DNA with double-stranded homologous ends and circular Golden-Gate assembly products. Meanwhile, the use of a restriction endonuclease to generate linear DNA with double-stranded homologous ends increases the efficiency of chemical and electrotransformation of Saccharomyces cerevisiae. Importantly, we also optimized the process such that both RCA and endonuclease treatment occur efficiently in the same buffer, streamlining the workflow and reducing product loss through purification steps. We expect that our approach could have utility beyond E. coli and S. cerevisiae and be applicable to areas such as directed evolution, genome engineering, and the manipulation of alternative organisms with even poorer transformation efficiencies.

DOI: https://doi.org/10.1021/cbe.4c00115
Chem Bio Eng. 2025 Feb 27. 2(2): 71-86

Nucleic Acid Framework-Enabled Spatial Organization for Biological Applications.

Rui Zhang, Xiaolei Zuo, Fangfei Yin.

Nucleic acid frameworks (NAFs) are artificially prepared from natural nucleic acids with a precise size and structure. DNA origami exhibits controllable 2D lamellar structure and thus is easily used to construct 3D structures with different morphologies. Tetrahedral DNA nanostructures (TDNs) are prepared with four DNA strands that hybridize to each other with a tetrahedral structure. Here we summarize molecular spatial organization with DNA origami and TDNs as models for 2D- and 3D-recombinations, discuss NAF-based biomimicking of proteins and biomembranes, and introduce the identification probes, functional groups, and intercalators for biosensing, bioimaging, and nanomedicine therapy. NAFs are also extended to applications to guide the formation of inorganic nanoparticles with precise size and structure. Thus, the NAFs exhibit special organization, are easy to functionalize, and are becoming an important platform for interdisciplinary study and applications, such as nanotechnology, biochemistry, synthetic biology, and nanomedicine.

DOI: https://doi.org/10.1021/cbe.4c00164
Macromolecules. 2025 Feb 25. 58(4): 2046-2052

Biomimetic Chemocatalytic Cascades-A New Strategy for Molecular Design of Degradable Polymer Systems.

Bin Tan, John R Dorgan.

Biological systems often involve cascading molecular signals; for example, blood coagulation involves a cascade of serial and parallel reactions catalyzed by enzymes. The present study draws inspiration from such complex biological systems to demonstrate, through a simple example, the purposeful design of a cascade system that enables control over polymer degradation kinetics. Micron size fibers of polylactide (PLA), cellulose acetate (CA), and their mixtures are subjected to hydrolysis at varying temperatures. Cleavage of the PLA produces an organic acid functional group that catalyzes the CA hydrolysis, thus demonstrating the use of synthetic molecular signaling. Furthermore, the presence of CA inhibits the degradation of PLA thereby demonstrating molecular feedback, another hallmark of biological molecular cascades. The parallel reaction cascade causes the hydrolysis rate constant for CA to increase 3.1 times compared to CA alone (from 5.7 × 10-4 to 1.78 × 10-3 L2 mol-2 h-1 at 125 °C); furthermore, due to molecular feedback, the hydrolysis rate constant for PLA decreases by 21% (from 2.40 × 10-3 to 1.90 × 10-3 L2 mol-2 h-1). The results demonstrate that synthetic signaling enables exquisitely tunable degradation kinetics. Technological applications of such purposely designed biomimetic systems are wide ranging and include the design of polymer systems for hydraulic fracturing, for biomedical applications, and for facilitating the recycling of mixed plastic wastes.

DOI: https://doi.org/10.1021/acs.macromol.4c02241
ACS Appl Mater Interfaces. 2025 Mar 01.

Sunlight-Driven Photomobile Polymer Materials Containing Push-Pull Azobenzene Moieties.

Toru Ube, Kuniaki Miyamoto, Seiji Kurihara, Tomiki Ikeda.

  Polymer materials that show macroscopic deformation upon irradiation with light are feasible as soft actuators. However, previous photomobile systems typically required artificial light sources for actuation. Herein, we develop photomobile polymer materials that can be deformed by natural sunlight. Azobenzenes functionalized with electron-donating and -withdrawing groups (push-pull azobenzenes) show trans-cis photoisomerization upon irradiation with sunlight and cis-trans thermal back isomerization after the cessation of irradiation. Push-pull azobenzenes are incorporated into crosslinked liquid-crystalline polymers, in which azobenzene moieties are uniaxially aligned. Upon irradiation with simulated sunlight, the polymer films exhibit bending toward the incident light and revert to the initial shapes after the discontinuation of irradiation. The time scales of the macroscopic bending and unbending are consistent with those of photoisomerization behaviors of azobenzene moieties. This result indicates that macroscopic deformation is induced through photochemical processes rather than photothermal processes. The development of sunlight-driven photomobile polymer materials would lead to the creation of autonomous and ecofriendly photoresponsive systems without the need for artificial optical elements, such as light sources, lenses, and filters.

Keywords:  azobenzene; liquid crystals; photoactuation; photochromism; stimuli-responsive polymers

DOI:  https://doi.org/10.1021/acsami.4c21123
STAR Protoc. 2025 Mar 05. pii: S2666-1667(25)00081-4. [Epub ahead of print]6(1): 103675

Protocol to modulate SUMOylation of a specific protein in budding yeast using chemical genetic approaches.

Emily Gutierrez-Morton, Yanchang Wang.

  SUMOylation (small ubiquitin-like modifier) is a ubiquitous and highly dynamic posttranslational modification. Here, we present a protocol to alter the local SUMOylation landscape of target proteins in budding yeast Saccharomyces cerevisiae using chemical genetic tools. We describe steps for recruiting SUMO enzymes (Ulp1PD or Ubc9) to GFP-tagged proteins using GBP (GFP-binding protein)-fusion proteins. We then detail procedures for inducing SUMO conjugation/deconjugation and the subsequent SUMOylation analysis. For complete details on the use and execution of this protocol, please refer to Gutierrez-Morton et al.1.

Keywords:  Cell Biology; Genetics; Molecular Biology

DOI:  https://doi.org/10.1016/j.xpro.2025.103675
Small Methods. 2025 Mar 03. e2500100

Liquid Printing in Nanochitin Suspensions: Interfacial Nanoparticle Assembly Toward Volumetric Elements, Organic Electronics and Core-Shell Filaments.

Mahyar Panahi-Sarmad, Ahmadreza Ghaffarkhah, Lukas Alexander Bauman, Amin Babaei-Ghazvini, Seyyed Alireza Hashemi, Bishnu Acharya, Boxin Zhao, Mohammad Arjmand, Feng Jiang, Orlando J Rojas.

  A nanoparticle-nanoparticle assembly is introduced using electrostatic complexation to precisely control volumetric structuring at the water/alcohol interface. In this system, an aqueous graphene oxide (GO) ink interacts electrostatically with partially deacetylated chitin nanofibers (mChNF), modified with benzophenone and dispersed in 1-butanol, which serves as the external phase. Upon extrusion of the GO ink, a jammed interfacial network forms, stabilizing the printed patterns within the external suspension, which provides suitable viscoelasticity for support-free printing. This approach is further extended to inks incorporating metal-organic frameworks or cellulose nanoparticles, demonstrating the advantages of mChNF as a stabilizer. Additionally, by incorporating a conductive polymer, the inks can be tailored for programmable and conductive patterning, opening new opportunities in liquid electronics and reconfigurable systems. Finally, GO inks containing an anionic polyelectrolyte (sodium alginate) undergo osmosis-driven solidification, facilitating the demolding of high-fidelity 3D structures formed by the printed threads of struts. These structures exhibit coreshell morphologies and high mechanical strength (∼175 MPa at 4% strain). Overall, this liquid-in-liquid fabrication approach, enabled by the integration of mChNF in the external phase, unlocks new possibilities for the design of versatile and multifunctional materials.

Keywords:  core–shell filaments; high‐resolution extrusion; liquid‐in‐liquid printing; nanoparticle interfacial assembly; partially miscible interfaces

DOI:  https://doi.org/10.1002/smtd.202500100
Soft Matter. 2025 Mar 06.

Adhesion study at the interface of a PDMS-elastomer and borosilicate glass-slide: effect of modulus and thickness of the elastomer.

Susheel Kumar, Chiranjit Majhi, Krishnacharya Khare, Manjesh K Singh.

Adhesion control at the interface of two surfaces is crucial in many applications. Examples are the design of micro and nanodevices such as microfluidic devices, biochips, and electronic sensors. Adhesion at the interface of two materials can be controlled by various methods, such as chemical treatment on the surface of the materials, modification of the surface texture of the materials, and change of the mechanical properties of the materials. The main idea of this study is to control the adhesion by changing the mechanical properties (modulus) of the polydimethylsiloxane (PDMS) elastomer. We vary the modulus of the PDMS elastomer by changing mixing ratio (w/w) of the silicone elastomer base and its curing agent (SylgardTM 184, Dow Corning). Our study also includes the effect of the thickness of the PDMS elastomer sheet on its adhesion behavior. Adhesion measurements at the interface of the borosilicate glass slide and different PDMS elastomer specimens were performed using a wedge test. This method inserts a glass coverslip at the interface to create a wedge. We observe a significant decrease in the work of adhesion and an increase in equilibrium crack length with an increase in elastic-modulus and thickness of the PDMS elastomer samples. We present and discuss the effect of modulus and specimen-thickness on the adhesion behavior of the PDMS elastomer against a glass slide.

DOI: https://doi.org/10.1039/d4sm01249f
Biofabrication. 2025 Mar 03.

High-resolution bioprinting of complex bio-structures via engineering of the photopatterning approaches and adaptive segmentation.

Ceren Babayigit, Jorge Alfonso Tavares Negrete, Rahim Esfandyarpour, Ozdal Boyraz.

  Digital Light Processing (DLP) technology has significantly advanced various applications, including 3D bioprinting, through its precision and speed in creating detailed structures. While traditional DLP systems rely on light-emitting diodes (LEDs), their limited power spectral density, high etendue, and spectral inefficiency constrain their performance in resolution, dynamic range, printing time, and cell viability. This study proposes and evaluates a dual-laser DLP system to overcome these limitations and enhance bioprinting performance. The proposed dual-laser system resulted in a twofold increase in resolution and a twelvefold reduction in printing time compared to the LED system. The system's capability was evaluated by printing three distinct designs, achieving a maximum percentage error of 1.16% and a minimum of 0.02% in accurately reproducing complex structures. Further, the impact of exposure times (10-30 s) and light intensities (0.044-0.11 mW/mm2) on the viability and morphology of 3T3 fibroblasts in GelMA and GelMA-PEGDA hydrogels is assessed. The findings reveal a clear relationship between longer exposure times and reduced cell viability. On day 7, samples exposed for extended periods exhibited the lowest metabolic activity and cell density, with differences of ~40% between treatments. However, all samples show recovery by day 7, with GelMA samples exhibiting up to a sixfold increase in metabolic activity and GelMA-PEGDA samples showing up to a twofold increase. In contrast, light intensity variations had a lesser effect, with a maximum variation of 15% in cell viability. We introduced a segmented printing method to mitigate over-crosslinking and enhance the dynamic range, utilizing an adaptive segmentation control strategy. This method, demonstrated by printing a bronchial model with a 14.43x compression ratio, improved resolution and maintained cell viability up to 90% for GelMA and 85% for GelMA-PEGDA during 7 days of culture. The proposed dual-laser system and adaptive segmentation method were confirmed through successful prints with diverse bio-inks and complex structures, underscoring its advantages over traditional LED systems in advancing 3D bioprinting.

Keywords:  3D Bioprinting; Adaptive Segmentation; Additive manufacturing; Bronchial-tree bioprinting; Digital light processing; Photopolymerization

DOI:  https://doi.org/10.1088/1758-5090/adbc22
PNAS Nexus. 2025 Feb;4(2): pgaf054

AI-enabled manufacturing process discovery.

D Quispe, D Kozjek, M Mozaffar, T Xue, J Cao.

  Discovering manufacturing processes has been largely experienced-based. We propose a shift to a systematic approach driven by dependencies between energy inputs and performance outputs. Uncovering these dependencies across diverse process classes requires a universal language that characterizes process inputs and performances. Traditional manufacturing languages, with their individualized syntax and terminology, hinder the characterization across varying length scales and energy inputs. To enable the evaluation of process dependencies, we propose a broad manufacturing language that facilitates the characterization of diverse process classes, which include energy inputs, tool-material interactions, material compatibility, and performance outputs. We analyze the relationships between these characteristics by constructing a dataset of over 50 process classes, which we use to train a variational autoencoder (VAE) model. This generative model encodes our dataset into a 2D latent space, where we can explore, select, and generate processes based on desired performances and retrieve the corresponding process characteristics. After verifying the dependencies derived from the VAE model match with existing knowledge on manufacturing processes, we demonstrate the usefulness of using the model to discover new potential manufacturing processes through three illustrative cases.

Keywords:  data-driven modeling; deep learning; manufacturing; variational autoencoder

DOI:  https://doi.org/10.1093/pnasnexus/pgaf054
Small. 2025 Mar 03. e2411506

Hybrid Ultrathin Gold Nanowire Gels: Formation and Mechanical Properties.

Yannic Curto, Srishti Arora, Bart-Jan Niebuur, Lola González-García, Tobias Kraus.

  This report is about the chemical formation of gels from ultrathin gold nanowires (AuNWs) and the gels' properties. An excess of triphenylphosphine (PPh3) initiated the gelation of AuNWs with core diameters below 2 nm and an oleylamine (OAm) ligand shell dispersed in cyclohexane. The ligand exchange of OAm by PPh3 changes the AuNW-solvent interactions and leads to phase separation of the solvent to form a macroscopic gel. Small angle X-ray scattering and transmission electron microscopy indicate that hexagonal bundles in the original dispersion are dispersed, and the released nanowires entangle. Rheological analyses indicate that the resulting gel is stabilized both by physical entanglement and crosslinking of AuNWs by Van der Waals and π-π interactions. Chemically formed AuNW gels have solid-like properties and crosslinks that distinguish them from highly concentrated non-crosslinked AuNW dispersions. The AuNW gel properties can be tuned via the Au:PPh3 ratio, where smaller ratios led to stiffer gels with higher storage moduli.

Keywords:  SAXS; colloidal gel; rheology; ultrathin gold nanowires

DOI:  https://doi.org/10.1002/smll.202411506
Adv Healthc Mater. 2025 Mar 06. e2403800

In Situ-Gelling Antimicrobial Poly(oligoethylene glycol methacrylate)-Based Hydrogels Integrating Bound Quaternary Ammonia Compounds and Antibiotic Functionalities for Effective Infected Wound Healing.

Fei Xu, Evelyn Cudmore, Sadru-Dean Walji, Lei Zhang, Meghan Kostashuk, Isabella Jun, Gurpreet Randhawa, Zhicheng Pan, Todd Hoare.

  In situ-gelling antibacterial hydrogels are reported in which two antibacterial entities (quaternary ammonium (QA) groups and the antibiotic ciprofloxacin (CIP)) are tethered to a single precursor based on the anti-fouling polymer poly(oligoethylene glycol methacrylate) (POEGMA). Synergism between the QA and CIP tethers is demonstrated to enable broad-spectrum killing and/or disinfection of both gram-positive and gram-negative bacteria both in vitro and in vivo while also supporting improved functional recovery of uninjured skin morphology. Coupled with the suitable mechanics, swelling capacity, and stability of the gels, the multi-mechanism antibacterial properties of the hydrogels offer promise for treating or preventing infections of burn wounds.

Keywords:  anti‐bacterial materials; hydrogels; in situ gelation; synergy; wound dressings

DOI:  https://doi.org/10.1002/adhm.202403800
Biochemistry. 2025 Mar 05.

Controlled Enzyme Cargo Loading in Engineered Bacterial Microcompartment Shells.

Nicholas M Tefft, Yali Wang, Alexander Jussupow, Michael Feig, Michaela A TerAvest.

Bacterial microcompartments (BMCs) are nanometer-scale organelles with a protein-based shell that serve to colocalize and encapsulate metabolic enzymes. They may provide a range of benefits to improve pathway catalysis, including substrate channeling and selective permeability. Several groups are working toward using BMC shells as a platform for enhancing engineered metabolic pathways. The microcompartment shell of Haliangium ochraceum (HO) has emerged as a versatile and modular shell system that can be expressed and assembled outside its native host and with non-native cargo. Further, the HO shell has been modified to use the engineered protein conjugation system SpyCatcher-SpyTag for non-native cargo loading. Here, we used a model enzyme, triose phosphate isomerase (Tpi), to study non-native cargo loading into four HO shell variants and begin to understand maximal shell loading levels. We also measured activity of Tpi encapsulated in the HO shell variants and found that activity was determined by the amount of cargo loaded and was not strongly impacted by the predicted permeability of the shell variant to large molecules. All shell variants tested could be used to generate active, Tpi-loaded versions, but the simplest variants assembled most robustly. We propose that the simple variant is the most promising for continued development as a metabolic engineering platform.

DOI: https://doi.org/10.1021/acs.biochem.4c00709
Soft Matter. 2025 Mar 03.

Micropores can enhance the intrinsic fracture energy of hydrogels.

Puyu Cao, Bin Chen, Yi Cao, Huajian Gao.

Hydrogels, a class of soft materials composed of a polymer chain network, are widely known to be prone to fatigue failure. To understand the underlying mechanisms, we simulate polymer scission and fatigue initiation in the vicinity of a crack tip within a two-dimensional polymer network. For a network without pores, our findings reveal that polymer scission can occur across multiple layers of chains, rather than just a single layer as assumed in the classical Lake-Thomas theory, consistent with previous studies. In contrast, for a network with a high density of micropores, our results demonstrate that the pores can substantially enhance the intrinsic fracture energy of the network in direct proportion to the pore size. This enhancement is attributed to pore-pore interactions, which lead to a relatively uniform distribution of cohesive energy ahead of the crack tip. Our model suggests that incorporating micropores could be a promising strategy for improving the intrinsic fracture energy of hydrogels and that natural porous tissues may have evolved to achieve enhanced fatigue resistance.

DOI: https://doi.org/10.1039/d4sm00973h
Lab Chip. 2025 Mar 03.

A microfluidic platform for extraction and analysis of bacterial genomic DNA.

Alex Joesaar, Martin Holub, Leander Lutze, Marco Emanuele, Jacob Kerssemakers, Martin Pabst, Cees Dekker.

Bacterial cells organize their genomes into a compact hierarchical structure called the nucleoid. Studying the nucleoid in cells faces challenges because of the cellular complexity while in vitro assays have difficulty in handling the fragile megabase-scale DNA biopolymers that make up bacterial genomes. Here, we introduce a method that overcomes these limitations as we develop and use a microfluidic device for the sequential extraction, purification, and analysis of bacterial nucleoids in individual microchambers. Our approach avoids any transfer or pipetting of the fragile megabase-size genomes and thereby prevents their fragmentation. We show how the microfluidic system can be used to extract and analyze single chromosomes from B. subtilis cells. Upon on-chip lysis, the bacterial genome expands in size and DNA-binding proteins are flushed away. Subsequently, exogeneous proteins can be added to the trapped DNA via diffusion. We envision that integrated microfluidic platforms will become an essential tool for the bottom-up assembly of complex biomolecular systems such as artificial chromosomes.

DOI: https://doi.org/10.1039/d4lc00839a
bioRxiv. 2025 Feb 23. pii: 2025.02.22.639668. [Epub ahead of print]

Cell size regulation in bacteria: a tale of old regulators with new mechanisms.

Ezza Khan, Paola E Mera.

Proper function in a bacterial cell relies on intrinsic cell size regulation. The molecular mechanisms underlying how bacteria maintain their cell size remain unclear. The conserved regulator DnaA, the initiator of chromosome replication, is associated to size regulation by controlling the number of origins of replication ( oriC ) per cell. In this study, we identify and characterize a new mechanism in which DnaA modulates cell size independently of oriC -copy number. By altering the levels of DnaA without impacting chromosome replication, we demonstrate that DnaA's activity as a transcription factor can slow down cell elongation rate resulting in cells that are ∼20% smaller. We identify the peptidoglycan biosynthetic enzyme MurD as a key player of cell size regulation in Caulobacter crescentus and in the evolutionarily distant bacterium Escherichia coli . Collectively, our findings provide mechanistic insights to the complex regulation of cell size in bacteria.

DOI: https://doi.org/10.1101/2025.02.22.639668
Nat Commun. 2025 Mar 04. 16(1): 2168

Computation-aided designs enable developing auxotrophic metabolic sensors for wide-range glyoxylate and glycolate detection.

Enrico Orsi, Helena Schulz-Mirbach, Charles A R Cotton, Ari Satanowski, Henrik M Petri, Susanne L Arnold, Natalia Grabarczyk, Rutger Verbakel, Karsten S Jensen, Stefano Donati, Nicole Paczia, Timo Glatter, Andreas M Küffner, Tanguy Chotel, Farah Schillmüller, Alberto De Maria, Hai He, Steffen N Lindner, Elad Noor, Arren Bar-Even, Tobias J Erb, Pablo I Nikel.

Auxotrophic metabolic sensors (AMS) are microbial strains modified so that biomass formation correlates with the availability of specific metabolites. These sensors are essential for bioengineering (e.g., in growth-coupled designs) but creating them is often a time-consuming and low-throughput process that can be streamlined by in silico analysis. Here, we present a systematic workflow for designing, implementing, and testing versatile AMS based on Escherichia coli. Glyoxylate, a key metabolite in (synthetic) CO2 fixation and carbon-conserving pathways, served as the test analyte. Through iterative screening of a compact metabolic model, we identify non-trivial growth-coupled designs that result in six AMS with a wide sensitivity range for glyoxylate, spanning three orders of magnitude in the detected analyte concentration. We further adapt these E. coli AMS for sensing glycolate and demonstrate their utility in both pathway engineering (testing a key metabolic module for carbon assimilation via glyoxylate) and environmental monitoring (quantifying glycolate produced by photosynthetic microalgae). Adapting this workflow to the sensing of different metabolites could facilitate the design and implementation of AMS for diverse biotechnological applications.

DOI: https://doi.org/10.1038/s41467-025-57407-3
Phys Life Rev. 2025 Feb 28. pii: S1571-0645(25)00020-X. [Epub ahead of print]53 91-116

Viscoelastic mechanics of living cells.

Hui Zhou, Ruye Liu, Yizhou Xu, Jierui Fan, Xinyue Liu, Longquan Chen, Qiang Wei.

  In cell mechanotransduction, cells respond to external forces or to perceive mechanical properties of their supporting substrates by remodeling themselves. This ability is endowed by modulating cells' viscoelastic properties, which dominates over various complex cellular processes. The viscoelasticity of living cells, a concept adapted from rheology, exhibits substantially spatial and temporal variability. This review aims not only to discuss the rheological properties of cells but also to clarify the complexity of cellular rheology, emphasizing its dependence on both the size scales and time scales of the measurements. Like typical viscoelastic materials, the storage and loss moduli of cells often exhibit robust power-law rheological characteristics with respect to loading frequency. This intrinsic feature is consistent across cell types and is attributed to internal structures, such as cytoskeleton, cortex, cytoplasm and nucleus, all of which contribute to the complexity of cellular rheology. Moreover, the rheological properties of cells are dynamic and play a crucial role in various cellular and tissue functions. In this review, we focus on elucidating time- and size-dependent aspects of cell rheology, the origins of intrinsic rheological properties and how these properties adapt to cellular functions, with the goal of interpretation of rheology into the language of cell biology.

Keywords:  Adaptation; Power-law; Size-dependent; Timescale-dependent; Viscoelasticity

DOI:  https://doi.org/10.1016/j.plrev.2025.02.004
Commun Biol. 2025 Mar 06. 8(1): 340

One-step Cre-loxP organism creation by TAx9.

Martin Miguel Casco-Robles, Tomoki Echigoya, Takeaki Shimazaki, Yuri Murakami, Masaya Hirano, Fumiaki Maruo, Seiya Mizuno, Satoru Takahashi, Chikafumi Chiba.

The creation of organisms with Cre-loxP conditional gene recombination systems often faces challenges, particularly when creating the initial (F0) generation with both a Cre recombinase and a DNA site flanked by loxP elements (floxed site). The primary reason is that it is difficult to synthesize a single plasmid with both the Cre gene and the floxed site, since Cre-mediated recombination spontaneously occurs when the plasmid is amplified in Escherichia coli bacterial cells. Here, we introduce an artificial nucleic acid sequence TATATATATATATATATA, named TAx9, that enables the integration of both the Cre gene and the floxed site into a single plasmid. TAx9 effectively blocks spontaneous Cre-mediated recombination in E. coli cells. Using this system, we created an F0 generation of transgenic newts and CRISPR-Cas9 knock-in mice with tissue-specific Cre recombination triggered by tamoxifen. TAx9 technology will be a powerful strategy for creating organisms capable of conditional genetic modification in the F0 generation, accelerating various life science research by reducing the time and cost for ultimately establishing and maintaining lines of genetically modified organisms.

DOI: https://doi.org/10.1038/s42003-025-07759-9
Cell Struct Funct. 2025 Mar 05.

Capturing CDKs in Action: Live-Cell Biosensors Pioneer the New Frontiers in Cell Cycle Research.

Sachiya Nakashima, Aika Toyama, Hironori Sugiyama, Kazuhiro Aoki, Yuhei Goto.

  Cyclin-dependent kinases (CDKs) orchestrate cell cycle progression through precise temporal control of substrate phosphorylation. While traditional biochemical approaches and phosphoproteomics have provided valuable insights into CDK-mediated regulation, these methods require cell population analyses and cannot capture real-time dynamics in individual cells. The recent development of fluorescent biosensors has revolutionized our ability to monitor CDK activity in living cells with unprecedented temporal and spatial resolution. Here, we comprehensively review genetically encoded fluorescent biosensors for measuring CDK activity. The two major modes of action in CDK activity biosensors-FRET-based and translocation-based biosensors-enable researchers to select appropriate tools for their specific experimental objectives. These biosensors have revealed precise spatiotemporal CDK activity dynamics across diverse model systems, including yeast, cultured mammalian cells, worms, flies, frog egg extract, fish, and mice. Such technological advances are transforming our understanding of quantitative principles underlying cell cycle control and opening new avenues for investigating cell cycle regulation in various biological contexts.Key words: CDK, FRET, cell cycle, live imaging, biosensor.

Keywords:  CDK; FRET; biosensor; cell cycle; live imaging

DOI:  https://doi.org/10.1247/csf.25004
J Am Chem Soc. 2025 Mar 04.

Bacteria-Mediated Intracellular Radical Polymerizations.

Eleonora Ornati, Jules Perrard, Tobias A Hoffmann, Raissa Bonon, Nico Bruns.

Intracellular radical polymerizations allow for the direct bioorthogonal synthesis of various synthetic polymers within living cells, thereby providing a pathway to polymer-modified cells or the fermentative production of polymers. Here, we show that Escherichia coli cells can initiate the polymerization of various acrylamide, acrylic, and methacrylic monomers through an atom transfer radical reaction triggered by the activity of naturally occurring biomolecules within the bacterial cells. Intracellular radical polymerizations were confirmed by nuclear magnetic resonance spectroscopy, gel permeation chromatography of polymers extracted from the cells, and fluorescence labeling of the polymer directly inside the cells. The effect of polymerization on cell behavior and the response of the cells to polymerization was investigated through fluorescence microscopy and flow cytometry techniques, as well as metabolic and membrane integrity assays. The polymer synthesis and resulting products are cell-compatible, as indicated by the high viability of the polymerized cells. In cellulo synthesis of synthetic polymers containing fluorescent dyes was also achieved. These results not only enhance our understanding of the untapped potential of bacterial cells as living catalysts for polymer production but also reveal intracellular polymerization based on atom transfer radical polymerization initiators as a bioorthogonal tool for cell engineering and synthetic biology.

DOI: https://doi.org/10.1021/jacs.4c17257
ACS Nano. 2025 Mar 02.

Facile Synthesis of Silicone Oil-Based Ferrofluid: Toward Smart Materials and Soft Robots.

Leilei Chen, Hengao Yu, Jilan Yang, Jinzhuo Shi, Chun-He Li, Zijie Qu, Wendong Wang.

  Ferrofluids are stable colloidal dispersions of magnetic nanoparticles in carrier liquids. Their combination of magnetic and fluidic characteristics not only inspires fundamental inquiries into forms and functions of matter but also enables diverse applications ranging from sealants and coolants in mechanical devices to active components in smart materials and soft robots. Spurred by such fundamental and applied interests, a growing need for easy-to-synthesize, high-quality ferrofluid exists. Here, we report the facile synthesis and comprehensive characterization of a silicone oil-based ferrofluid that displays the characteristic surface instability of high-quality ferrofluids and demonstrate its functions in smart interfacial materials and soft robots. Silicone's chemical immiscibility with polar solvents and its biological inertness, when coupled with magnetic responsiveness and fluidic deformability, enable the manipulation of solid particles, gas bubbles, simple and complex liquids, as well as micro-organisms. We envision that the silicone oil-based ferrofluid will find applications in diverse areas, including magnetic digital microfluidics, multifunctional materials, and small-scale robotics.

Keywords:  ferrofluid; magnetic materials; programmable materials; self-assembly; smart materials; soft robots

DOI:  https://doi.org/10.1021/acsnano.4c16689
J Nanobiotechnology. 2025 Mar 07. 23(1): 187

Overcoming ice: cutting-edge materials and advanced strategies for effective cryopreservation of biosample.

Miaorong Huang, Minhua Hu, Gengyuan Cai, Hengxi Wei, Sixiu Huang, Enqin Zheng, Zhenfang Wu.

  Cryopreservation techniques have been widely used, especially in biomedical applications and preservation of germplasm resources. Ideally, biological materials would maintain functional integrity as well as a normal structure and can be recovered when needed. However, this tool does not work all the time. Ice formation and growth are the key challenges. The other major reason is that the cryoprotective agents (CPAs) currently used do not meet these needs and are always accompanied by their cytotoxicity. A comprehensive and synergistic approach that focuses on the overall frozen biological system is crucial for the evolution of cryopreservation methods. In this review, we first summarize the fundamental damage mechanisms during cryopreservation, as well as common cryoprotectants and their limitations. Next, we discuss materials that interact with ice to improve cryopreservation outcomes. We evaluated natural and synthetic materials, including sugars and polymers, AFPs and mimics, ice nucleators, and hydrogels. In addition, biochemical regulation, which enhances the tolerance of biosamples to cryopreservation-induced stresses, was also mentioned. Nanotechnology, cell encapsulation, cryomesh, and isochoric freezing, such scalable approaches, are further discussed for cryopreservation. Finally, future research directions in this field for efficient cryopreservation are proposed. We emphasized the need for multidisciplinary progress to address these challenges. The combination of cryobiology mechanisms with technologies, such as synthetic biology, nanotechnology, microfluidics, and 3D bioprinting, is highlighted.

Keywords:  3D bioprinting; Antioxidant; Cryopreservation; Hydrogels; Microfluidics; Nanotechnology

DOI:  https://doi.org/10.1186/s12951-025-03265-6
NPJ Metab Health Dis. 2025 ;3(1): 7

Regulation of gene expression through protein-metabolite interactions.

Maximilian Hornisch, Ilaria Piazza.

  Organisms have to adapt to changes in their environment. Cellular adaptation requires sensing, signalling and ultimately the activation of cellular programs. Metabolites are environmental signals that are sensed by proteins, such as metabolic enzymes, protein kinases and nuclear receptors. Recent studies have discovered novel metabolite sensors that function as gene regulatory proteins such as chromatin associated factors or RNA binding proteins. Due to their function in regulating gene expression, metabolite-induced allosteric control of these proteins facilitates a crosstalk between metabolism and gene expression. Here we discuss the direct control of gene regulatory processes by metabolites and recent progresses that expand our abilities to systematically characterize metabolite-protein interaction networks. Obtaining a profound map of such networks is of great interest for aiding metabolic disease treatment and drug target identification.

Keywords:  Biochemistry; Chemical biology; Systems biology

DOI:  https://doi.org/10.1038/s44324-024-00047-w
Small Methods. 2025 Mar 03. e2401969

Synthetic Biology-Based Strategy for Nanomedicine Design.

Xiangyun Zhang, Qiannan Duan, Jie Zhuang, Xinglu Huang.

  Nanomedicines have demonstrated significant potential in disease diagnosis and therapy, revolutionizing traditional drug development patterns. Recently, inspired by both natural and engineering principles, synthetic biology integrates the complexity of biological systems with the precision of engineering to design and create novel biological components, devices, and systems. This convergence of synthetic biology and nanomedicine has led to the emergence of a new concept: synthetic biological nanomedicine. Unlike traditional or biomimetic nanomedicines, synthetic biological nanomedicines are designed using gene engineering-based strategies. In this Perspective, the foundational concepts of synthetic biological nanomedicine are introduced and its relationship to, and differences from, traditional and biomimetic nanomedicine are explored. Drawing from synthetic biology, synthetic biological nanomedicine also incorporates two main approaches: top-down and bottom-up strategies. The latest advancements in the application of synthetic biology to nanomedicine are reviewed, these developments are categorized according to the aforementioned strategies, and a discussion of the potential advantages and challenges associated with utilizing synthetic biology in nanomedicine development is concluded.

Keywords:  bottom‐up strategy; genetic engineering; nanomedicines; synthetic biology; top‐down strategy

DOI:  https://doi.org/10.1002/smtd.202401969
ACS Appl Bio Mater. 2025 Mar 07.

From Unprintable Peptidic Gel to Unstoppable: Transforming Diphenylalanine Peptide (Fmoc-FF) Nanowires and Cellulose Nanofibrils into a High-Performance Biobased Gel for 3D Printing.

Feras Dalloul, J Benedikt Mietner, Dhanya Raveendran, Shouzheng Chen, Enguerrand Barba, Dennis M J Möck, Fabio Hubel, Benedikt Sochor, Sarathlal Koyiloth Vayalil, Linnea Hesse, Andrea Olbrich, Jörn Appelt, Peter Müller-Buschbaum, Stephan V Roth, Julien R G Navarro.

  The growing interest in gel-based additive manufacturing, also known as three-dimensional (3D) gel-printing technology, for research underscores the crucial need to develop robust biobased materials with excellent printing quality and reproducibility. The main focus of this study is to prepare and characterize some composite gels obtained with a low-molecular-weight gelling (LMWG) peptide called Fmoc-diphenylalanine (Fmoc-FF) and two types of cellulose nanofibrils (CNFs). The so-called Fmoc-FF peptide has the ability to self-assemble into a nanowire shape and therefore create an organized network that induces the formation of a gel. Despite their ease of preparation and potential use in biological systems, unfortunately, those Fmoc-FF nanowire gel systems cannot be 3D printed due to the high stiffness of the gel. For this reason, this study focuses on composite materials made of cellulose nanofibrils and Fmoc-FF nanowires, with the main objective being that the composite gels will be suitable for 3D printing applications. Two types of cellulose nanofibrils are employed in this study: (1) unmodified pristine cellulose nanofibrils (uCNF) and (2) chemically modified cellulose nanofibrils, which ones have been grafted with polymers containing the Fmoc unit on their backbone (CNF-g-Fmoc). The obtained products were characterized through solid-state cross-polarization magic angle-spinning 1H NMR and confocal laser scanning microscopy. Within these two CNF structures, two composite gels were produced: uCNF/Fmoc-FF and CNF-g-Fmoc/Fmoc-FF. The mechanical properties and printability of the composites are assessed using rheology and challenging 3D object printing. With the addition of water, different properties of the gels were observed. In this instance, CNF-g-Fmoc/Fmoc-FF (c = 5.1%) was selected as the most suitable option within this product range. For the composite bearing uCNF, exceptional print quality and mechanical properties are achieved with the CNF/Fmoc-FF gel (c = 5.1%). The structures are characterized by using field emission scanning electron microscopy (FESEM) and small-angle X-ray scattering (SAXS) measurements.

Keywords:  3D gel printing; Fmoc-FF; cellulose nanofibrils (CNF); direct ink writing (DIW); nanocellulose; single electron transfer living radical polymerization (SET-LRP)

DOI:  https://doi.org/10.1021/acsabm.4c01803
Macromolecules. 2025 Feb 25. 58(4): 2073-2084

Presenting Antimicrobial Peptides on Poly(ethylene glycol): Star-Shaped vs Comb-Like Architectures.

Zixian Cui, Elliot A Brna, Matthew A Crawford, Puthayalai Treerat, Mobina Alimadad, Molly A Hughes, Rachel A Letteri.

Conjugating antimicrobial peptides (AMPs) to nonlinear polymers is a promising strategy to overcome the translational challenges of AMPs toward treating infections caused by antibiotic-resistant bacteria. Nonlinear polymers, and therefore conjugates, can be prepared with various architectures (e.g., star-shaped, comb-like, hyperbranched, etc.), however, the effects of polymer architecture on antimicrobial performance and related properties, like size and morphology in solution and secondary structure, are not yet well-understood. Here, we compare conjugates of the human chemokine-derived AMP stapled P9 with poly(ethylene glycol) (PEG) prepared in two of the major nonlinear architectures: star-shaped and comb-like. At comparable molecular weights and compositions (peptide wt %), comb-like conjugates afford increased helicity, solubility, antimicrobial activity, and proteolytic stability compared to star-shaped analogs. We then leveraged the expansive design space of comb-like architectures to prepare conjugates with different backbone lengths and PEG side chain lengths, with shorter PEG side chains leading to increased helicity, yet potentially less shielding from proteolytic degradation and the longest backbone lengths furnishing the most potent antimicrobial activity. Both comb-like and star-shaped conjugates display high zeta potential, indicating that the cationic AMPs were accessible for electrostatic interactions with bacterial membranes. Yet, the comb-like conjugates showed a higher fraction of unimolecular structures indicative of a lower propensity for supramolecular assembly that could be encumbering the desired AMP-bacteria interactions in the star-shaped conjugates. Together, our work shows comb-like AMP-polymer conjugates to outperform analogous star-shaped conjugates, while adding design flexibility to access an expansive range of monomer chemistries, monomer distributions, and backbone lengths to modulate performance-determining properties and ultimately furnish an effective suite of AMP-polymer materials as alternatives to conventional antibiotics for combatting bacterial infections.

DOI: https://doi.org/10.1021/acs.macromol.4c02762
Small. 2025 Mar 04. e2500531

Site-Specific Polymer-Protein-Polymer Conjugates for the Preparation of Dual Responsive Multilayer Nanoparticles.

Melina I Feldhof, Simon Walber, Sandro Sperzel, Susanne Boye, Ulla I M Gerling-Driessen, Laura Hartmann.

  Protein-polymer-based materials demonstrate high potential in advanced applications. However, controlled combinations of multiple proteins and polymers to obtain multimaterial systems is limited due to the complexity of retaining protein structure and function and achieving high structural control for the polymers simultaneously. Here, the first combination of a rebridging agent and thiol-induced, light-activated controlled radical polymerization (TIRP) is introduced to directly enable site-specific conjugation of two different polymers to native proteins. Specifically, poly(N-isopropyacrylamide) (pNIPAM) is attached to bovine serum albumin (BSA), followed by incorporation of a new rebridging agent, and initiating a second TIRP to introduce a glycopolymer, giving highly defined pNIPAM-BSA-glycopolymer conjugates. Above the lower critical solution temperature (LCST), nanoparticles with a glycopolymer corona are formed. The addition of a glycan-specific lectin leads to the formation of a second protein corona and so-called multilayer nanoparticles. Depending on the sequence of stimuli, the particles can either undergo a step-wise or one-step disassembly. Furthermore, by controlling the ratio of binding/non-binding glycopolymers in the multilayer nanoparticles, either distinct nanoparticles or large clusters can be formed. Thus, dual-responsive multilayered polymer-protein nanoparticles are now accessible with controlled and programmable material properties such as assembly and disassembly while maintaining the protein's native structure and thus function.

Keywords:  dual responsive multilayer nanoparticles; ligand‐receptor interaction; polymer‐protein‐polymer conjugates; rebridging agent; thiol‐induced light activated controlled radical polymerization (TIRP)

DOI:  https://doi.org/10.1002/smll.202500531