Functional coating materials have found broad technological applications in diverse fields. Despite recent advances, few coating materials simultaneously achieve robustness and substrate independence while still retaining the capacity for genetically encodable functionalities. Here, we report Escherichia coli biofilm-inspired protein nanofiber coatings that simultaneously exhibit substrate independence, resistance to organic solvents, and programmable functionalities. The intrinsic surface adherence of CsgA amyloid proteins, along with a benign solution-based fabrication approach, facilitates forming nanofiber coatings on virtually any surface with varied compositions, sizes, shapes, and structures. In addition, the typical amyloid structures endow the nanofiber coatings with outstanding robustness. On the basis of their genetically engineerable functionality, our nanofiber coatings can also seamlessly participate in functionalization processes, including gold enhancement, diverse protein conjugations, and DNA binding, thus enabling a variety of proof-of-concept applications, including electronic devices, enzyme immobilization, and microfluidic bacterial sensors. We envision that our coatings can drive advances in electronics, biocatalysis, particle engineering, and biomedicine.
Abstract Marine diatoms construct their hierarchically ordered, three-dimensional (3D) external structures called frustules through precise biomineralization processes. Recapitulating the remarkable architectures and functions of diatom frustules in artificial materials is a major challenge that has important technological implications for hierarchically ordered composites. Here, we report the construction of highly ordered, mineralized composites based on fabrication of complex self-supporting porous structures—made of genetically engineered amyloid fusion proteins and the natural polysaccharide chitin—and performing in situ multiscale protein-mediated mineralization with diverse inorganic materials, including SiO2, TiO2 and Ga2O3. Subsequently, using sugar cubes as templates, we demonstrate that 3D fabricated porous structures can become colonized by engineered bacteria and can be functionalized with highly photoreactive minerals, thereby enabling co-localization of the photocatalytic units with a bacteria-based hydrogenase reaction for a successful semi-solid artificial photosynthesis system for hydrogen evolution. Our study thus highlights the power of coupling genetically engineered proteins and polysaccharides with biofabrication techniques to generate hierarchically organized mineralized porous structures inspired by nature.
There is an increasing trend of combining living cells with inorganic semiconductors to construct semi-artificial photosynthesis systems. Creating a robust and benign bio-abiotic interface is key to the success of such solar-to-chemical conversions but often faces a variety of challenges, including biocompatibility and the susceptibility of cell membrane to high-energy damage arising from direct interfacial contact. Here, we report living mineralized biofilms as an ultrastable and biocompatible bio-abiotic interface to implement single enzyme to whole-cell photocatalytic applications. These photocatalyst-mineralized biofilms exhibited efficient photoelectrical responses and were further exploited for diverse photocatalytic reaction systems including a whole-cell photocatalytic CO 2 reduction system enabled by the same biofilm-producing strain. Segregated from the extracellularly mineralized semiconductors, the bacteria remained alive even after 5 cycles of photocatalytic NADH regeneration reactions, and the biofilms could be easily regenerated. Our work thus demonstrates the construction of biocompatible interfaces using biofilm matrices and establishes proof of concept for future sustainable photocatalytic applications.
Proton conduction is essential in biological systems. Oxidative phosphorylation in mitochondria, proton pumping in bacteriorhodopsin and uncoupling membrane potentials by the antibiotic Gramicidin are examples. In these systems, H+ hop along chains of hydrogen bonds between water molecules and hydrophilic residues – proton wires. These wires also support the transport of OH− as proton holes. Discriminating between H+ and OH− transport has been elusive. Here, H+ and OH− transport is achieved in polysaccharide- based proton wires and devices. A H+- OH− junction with rectifying behaviour and H+-type and OH−-type complementary field effect transistors are demonstrated. We describe these devices with a model that relates H+ and OH− to electron and hole transport in semiconductors. In turn, the model developed for these devices may provide additional insights into proton conduction in biological systems.
Event Abstract Back to Event Engineering Bacillus subtilis Biofilms as Living Functional Materials Jiaofang Huang1, Chen Zhang1, Tianxin Zhao1, Ke Li1, Xinyu Wang1* and Chao Zhong1 1 ShanghaiTech University, School of Physical Science and Technology, China Introduction: Living functional biofilms are of keen interest for various applications [1],[2], as such materials system seamlessly integrates the tunable functional properties of amyloids with the environmental responsiveness and self-regeneration of living cells. Despite important advances [1],[2], such materials system has only been demonstrated in Escherichia coli, preventing their practical applications in medical fields. Furthermore, secretion of amyloid monomers with fused domains (>59 amino acids) in E. coli had been proved difficult due to the outer membrane feature of Gram-negative bacteria. Here we report a newly engineered living functional biofilm platform, based on Bacillus subtilis, a FDA (Food and Drug Administration)-approved and GRAS (Generally Recognized As Safe) Gram-positive bacteria species that can secrete amyloid fusion proteins with larger size (92 amino acids). We are leveraging this living functional biofilm platform for medical and environmental applications. Materials and Methods: 2.1 Strains and plasmids The strains used in this study were Bacillus subitlis NCBI 3610 cured strain DS2569 (3610 lacking pBS32) [3] and tasA mutant. Four fusion fragments were inserted into pHT01 vector respectively. 2.2 Biofilm formation The medium used for biofilm formation was MSgg with specific components listed in [4]. Congo Red indicator plates were MSgg agar containing 20 μg/mL Congo Red and 5 μg/mL Coomassie Brilliant Blue G. Media were solidified through addition of agar to 1.5%. Antibiotic concentrations were chloramphenicol (5 μg/mL) and Ampicillin (100 μg/mL). 2.3 Transmission Electron Microscopy (TEM) Sample preparation for TEM followed the protocol in 1. The samples were dried and examined in a JEOL 1200EX TEM at an accelerating voltage of 120 kV. Results and Discussion: Specifically, TasA, major protein component of B. subtilis biofilms [5], was genetically fused with a variety of functional domains, including TasA-Histag, TasA-Spytag, TasA-Mfp3 (Mussel foot proteins 3) and TasA-Mfp5 (Mussel foot protein 5) [6]. The engineered TasA fusion proteins can be secreted and extracellularly self-assemble into amyloid nanofibers with retained functions derived from the displayed protein/peptide proteins, as demonstrated in Congo Red (CR) (Figure 1A), TEM (Figure 1B), SEM (not shown) and other assays (not shown). Notably, the longest length of fusion peptides that could be secreted and adhered to biofilms reaches 92 amino acids, significantly outperforming the E. coli biofilm system. Figure 1 B. subtilis wild-type and tasA mutant with TasA-fusion expression strains grown on Congo Red (CR) plates (A), with morphology revealed by TEM (B), all scale bars are 1µm. Note: CR Assay indicates that wild-type and tasA mutant Bacillus strains with TasA-fusion expression plasmids all displayed enhanced red color compared with the tasA mutant with empty plasmid (A). TasA-fusion expression strains produced amyloid nanofibers based on TEM (B). Conclusions: Our work demonstrate that TasA nanofibers, major protein components of B. subtilis biofilms, could be genetically modified and secreted extracelluarly with additional functions introduced via peptide/protein fusion domains, thereby enabling a new living functional biofilm platform. Notably, the longest length of fusion peptides that could be secreted and adhered to biofilms reaches 92 amino acids, outperforming the E. coli biofilm system [2]. This work was supported in part by grant from the National Natural Science Foundation of China (NSFC) (31400042).References:[1] Chen, A.Y. et al. Synthesis and patterning of tunable multiscale materials with engineered cells. Nature materials 13, 515-523 (2014).[2] Nguyen, P.Q., Botyanszki, Z., Tay, P.K.R. & Joshi, N.S. Programmable biofilm-based materials from engineered curli nanofibres. Nature communications 5 (2014).[3] Konkol, M.A., Blair, K.M. & Kearns, D.B. Plasmid-encoded ComI inhibits competence in the ancestral 3610 strain of Bacillus subtilis. J Bacteriol 195, 4085-93 (2013).[4] Branda, S.S., Chu, F., Kearns, D.B., Losick, R. & Kolter, R. A major protein component of the Bacillus subtilis biofilm matrix. Mol Microbiol 59, 1229-38 (2006).[5] Romero, D., Aguilar, C., Losick, R. & Kolter, R. Amyloid fibers provide structural integrity to Bacillus subtilis biofilms. Proceedings of the National Academy of Sciences 107, 2230-2234 (2010).[6] Zhong, C. et al. Strong underwater adhesives made by self-assembling multi-protein nanofibres. Nature nanotechnology (2014). Keywords: self-assembly, biomaterial, protein, bioinerface Conference: 10th World Biomaterials Congress, Montréal, Canada, 17 May - 22 May, 2016. Presentation Type: Poster Topic: Protein-based biomaterials Citation: Huang J, Zhang C, Zhao T, Li K, Wang X and Zhong C (2016). Engineering Bacillus subtilis Biofilms as Living Functional Materials. Front. Bioeng. Biotechnol. Conference Abstract: 10th World Biomaterials Congress. doi: 10.3389/conf.FBIOE.2016.01.01399 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 27 Mar 2016; Published Online: 30 Mar 2016. * Correspondence: Dr. Xinyu Wang, ShanghaiTech University, School of Physical Science and Technology, Shanghai, China, wangxy@shanghaitech.edu.cn Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Jiaofang Huang Chen Zhang Tianxin Zhao Ke Li Xinyu Wang Chao Zhong Google Jiaofang Huang Chen Zhang Tianxin Zhao Ke Li Xinyu Wang Chao Zhong Google Scholar Jiaofang Huang Chen Zhang Tianxin Zhao Ke Li Xinyu Wang Chao Zhong PubMed Jiaofang Huang Chen Zhang Tianxin Zhao Ke Li Xinyu Wang Chao Zhong Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.
Nanoscale objects feature very large surface-area-to-volume ratios and are now understood as powerful tools for catalysis, but their nature as nanomaterials brings challenges including toxicity and nanomaterial pollution. Immobilization is considered a feasible strategy for addressing these limitations. Here, as a proof-of-concept for the immobilization of nanoscale catalysts in the extracellular matrix of bacterial biofilms, we genetically engineered amyloid monomers of the Escherichia coli curli nanofiber system that are secreted and can self-assemble and anchor nano-objects in a spatially precise manner. We demonstrated three scalable, tunable and reusable catalysis systems: biofilm-anchored gold nanoparticles to reduce nitro aromatic compounds such as the pollutant p-nitrophenol, biofilm-anchored hybrid Cd0.9Zn0.1S quantum dots and gold nanoparticles to degrade organic dyes and biofilm-anchored CdSeS@ZnS quantum dots in a semi-artificial photosynthesis system for hydrogen production. Our work demonstrates how the ability of biofilms to grow in scalable and complex spatial arrangements can be exploited for catalytic applications and clearly illustrates the design utility of segregating high-energy nano-objects from injury-prone cellular components by engineering anchoring points in an extracellular matrix.
Proton conduction is essential in biological systems. Oxidative phosphorylation in mitochondria, proton pumping in bacteriorhodopsin, and uncoupling membrane potentials by the antibiotic Gramicidin are examples. In these systems, H+ hop along chains of hydrogen bonds between water molecules and hydrophilic residues – proton wires. These wires also support the transport of OH− as proton holes. Discriminating between H+ and OH− transport has been elusive. Here, H+ and OH− transport is achieved in polysaccharide- based proton wires and devices. A H+- OH− junction with rectifying behaviour and H+-type and OH−-type complementary field effect transistors are demonstrated. We describe these devices with a model that relates H+ and OH− to electron and hole transport in semiconductors. In turn, the model developed for these devices may provide additional insights into proton conduction in biological systems.
Event Abstract Back to Event A genetic and modular design strategy towards multi-functional and self-assembling underwater adhesives Chao Zhong1*, Mengkui Cui1 and Bolin An1 1 ShanghaiTech University, School of Physical Science and Technology, China Underwater adhesives with tunable compositions and functions have many important applications in both biomedical and marine fields, but remain a big challenge in current adhesive techniques. In previous study, we demonstrated a genetic modular strategy for engineering underwater adhesives based on the fusion of mussel foot proteins (Mfps) from Mytilus galloprovincialis with CsgA, the major subunit of adhesive curli fibers from Escherichia coli [1]. However, these fibrous materials cannot satisfy all the requirements of ideal medical adhesives and their associated mechanism for underwater adhesion is also not fully understood. We utilize surface force apparatus (SFA) and atomic force microscopy (AFM) colloidal probe techniques, coupled with the genetic modular design strategy, to investigate the underwater adhesion mechanism of a variety of fusion proteins with tailor-designed protein domains. Furthermore, using ideal medical adhesives as blueprint, we leverage the genetic modular design strategy to further engineer biocompatible, injectable and ultra-strong adhesives based upon rational recombination of an amyloid-like gelation domain, cell adhesion moieties and mussel foot adhesive proteins. Collectively, this study will provide new insights into the adhesion mechanism of a new type of bio-inspired amyloid underwater adhesives and lay the foundation for engineering multi-compositional and multifunctional underwater adhesives. References:[1] C. Zhong, T. Gurry, A. Cheng, J. Downey, Z. T. Deng, M. Stultz, T. K. Lu. Strong Underwater Adhesives Made by Self-assembling Multi-protein Nanofibers. Nature Nanotechnology 2014, 10, 858-866. Keywords: self-assembly, Biomimetic, biomaterial, protein Conference: 10th World Biomaterials Congress, Montréal, Canada, 17 May - 22 May, 2016. Presentation Type: Poster Topic: Adhesive biomaterials Citation: Zhong C, Cui M and An B (2016). A genetic and modular design strategy towards multi-functional and self-assembling underwater adhesives. Front. Bioeng. Biotechnol. Conference Abstract: 10th World Biomaterials Congress. doi: 10.3389/conf.FBIOE.2016.01.01335 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 27 Mar 2016; Published Online: 30 Mar 2016. * Correspondence: Dr. Chao Zhong, ShanghaiTech University, School of Physical Science and Technology, Shanghai, China, Email1 Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Chao Zhong Mengkui Cui Bolin An Google Chao Zhong Mengkui Cui Bolin An Google Scholar Chao Zhong Mengkui Cui Bolin An PubMed Chao Zhong Mengkui Cui Bolin An Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.
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