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During the past decades, the importance of developing sustainable, carbon dioxide (CO 2 )-neutral and biodegradable alternatives to conventional plastic has become evident in the context of global pollution issues. Therefore, heterotrophic bacteria such as Cupriavidus sp. have been intensively explored for the synthesis of the biodegradable polymer polyhydroxybutyrate (PHB). PHB is also naturally produced by a variety of phototrophic cyanobacteria, which only need sunlight and CO 2, thereby allowing a CO 2 negative, eco-friendly synthesis of this polymer. However, a major drawback of the use of cyanobacteria is the need of a two-stage production process, since relevant amount of PHB synthesis only occurs after transferring the cultures to conditions of nitrogen starvation, which hinders continuous, large-scale production. This study aimed at generating, by means of genetic engineering, a cyanobacterium that continuously produces PHB in large amounts. We choose a genetically amenable filamentous cyanobacterium of the genus Nostoc sp., which is a diazotrophic cyanobacterium, capable of atmospheric nitrogen (N 2 ) fixation but naturally does not produce PHB. We transformed this Nostoc strain with various constructs containing the constitutive promotor P psbA and the PHB synthesis operon phaC1AB from Cupriavidus necator H16. In fact, while the transformants initially produced PHB, the PHB-producing strains rapidly lost cell viability. Therefore, we next attempted further optimization of the biosynthetic gene cluster. The PHB operon was expanded with phasin gene phaP1 from Cupriavidus necator H16 in combination with the native intergenic region of apcBA from Nostoc sp. 7120. Finally, we succeeded in stabilized PHB production, whilst simultaneously avoiding decreasing cell viability. In conclusion, the recombinant Nostoc strain constructed in the present work constitutes the first example of a continuous and stable PHB production platform in cyanobacteria, which has been decoupled from nitrogen starvation and, hence, harbours great potential for sustainable, industrial PHB production.

The primordial cyanobacteria were responsible for developing oxygenic photosynthesis early in evolution. In the pathways of fixed carbon allocation, the co-factor-independent phosphoglycerate mutase (iPGAM) plays a crucial role by directing the first CO 2 fixation product, 3-phosphoglycerate, toward central anabolic glycolytic-derived pathways. This work reveals a distinct evolution of iPGAM within oxygenic photosynthetic organisms. We have identified two specific segments in cyanobacterial iPGAMs that affect the control of iPGAM activity through its specific interactor protein PirC. This understanding of iPGAM has allowed us to engineer cyanobacterial strains with altered carbon fluxes. Since cyanobacteria can directly convert CO 2 into valuable products, our results demonstrate a novel approach for developing a chassis for biotechnical use.

Large‐scale production of polyhydroxybutyrate (PHB), a biodegradable bioplastic, using genetically engineered cyanobacteria offers a sustainable alternative to petrochemical‐derived plastics. However, monoculture‐based phototrophic systems face major limitations, such as poor resilience in large‐scale reactors, hindering industrial upscaling. To address these challenges, we replaced the native cyanobacterium of a natural microbial consortium with a genetically engineered Synechocystis strain optimised for PHB production, establishing what we define a hybrid photosynthetic microbiome. This new community preserved the ecological structure and stability of the original microbiome while gaining synthetic production capacity. Compared to the axenic strain, the hybrid system exhibited enhanced robustness under abiotic stress, including light and temperature fluctuations, and improved tolerance to operational instability. These features made it suitable for upscaling and application in non‐sterile environments. The hybrid microbiome sustained PHB production in scaled photobioreactors, reaching up to 32% PHB per cell dry weight (CDW) equal to ~230 mg L−1 under fully photoautotrophic conditions. Production was also achieved under dark conditions with acetate supplementation, highlighting the system's metabolic flexibility. This work demonstrates the successful integration of an engineered phototroph into a stable native microbiome, positioning hybrid communities as powerful platform for industrial biotechnology. We developed a hybrid photosynthetic microbiome, in which the native cyanobacterium of a natural microbial consortium was replaced with a genetically engineered Synechocystis strain optimised for polyhydroxybutyrate (PHB) production. The hybrid system demonstrated improved tolerance to abiotic stresses, making it suitable for upscaling.

Large-scale production of polyhydroxybutyrate (PHB), a biodegradable bioplastic, using genetically engineered cyanobacteria offers a sustainable alternative to petrochemical-derived plastics. However, monoculture-based phototrophic systems face major limitations, such poor resilience in large-scale reactors, hindering industrial upscaling. To address these challenges, we established a hybrid photosynthetic microbiome by replacing the native cyanobacterium of a natural microbial consortium with a genetically engineered Synechocystis strain optimized for PHB production. This new community retained the ecological structure and stability of the original microbiome while gaining synthetic production capacity. Compared to the axenic strain, the hybrid system exhibited enhanced robustness under abiotic stress, including light and temperature fluctuations, and improved tolerance to operational instability. These features made it suitable for upscaling and application in non-sterile environments. The hybrid microbiome sustained PHB production in scaled photobioreactors, reaching up to 32% PHB per cell dry weight (CDW) equal to ∼230 mg L-1 under fully photoautotrophic conditions. Production was also achieved under dark conditions with acetate supplementation, highlighting the system’s metabolic flexibility. This work demonstrates the successful integration of an engineered phototroph into a stable native microbiome, positioning hybrid communities as powerful platform for industrial biotechnology.

During the past decades, the importance of developing sustainable, carbon dioxide (CO2)-neutral and biodegradable alternatives to conventional plastic has become evident in the context of global pollution issues. Therefore, heterotrophic bacteria such as Cupriavidus sp. have been intensively explored for the synthesis of the biodegradable polymer polyhydroxybutyrate (PHB). PHB is also naturally produced by a variety of phototrophic cyanobacteria, which only need sunlight and CO2, thereby allowing a CO2 negative, eco-friendly synthesis of this polymer. However, a major drawback of the use of cyanobacteria is the need of a two-stage production process, since relevant amount of PHB synthesis only occurs after transferring the cultures to conditions of nitrogen starvation, which hinders continuous, large-scale production. This study aimed at generating, by means of genetic engineering, a cyanobacterium that continuously produces PHB in large amounts. We choose a genetically amenable filamentous cyanobacterium of the genus Nostoc sp., which is a diazotrophic cyanobacterium, capable of atmospheric nitrogen (N2) fixation but naturally does not produce PHB. We transformed this Nostoc strain with various constructs containing the PHB synthesis operon from Cupriavidus necator H16. In fact, while the transformants initially produced PHB, the PHB-producing strains rapidly lost cell viability. Therefore, we next attempted further optimization of the biosynthetic gene cluster. Finally, we succeeded in stabilized PHB production, whilst simultaneously avoiding decreasing cell viability. In conclusion, the recombinant Nostoc strain constructed in the present work constitutes the first example of a continuous and stable PHB production platform in cyanobacteria, which has been decoupled from nitrogen starvation and, hence, harbours great potential for sustainable, industrial PHB production.

The 2,3-bisphosphoglycerate-independent phosphoglycerate mutase (iPGAM) has been identified as a crucial regulating key point in the carbon storage metabolism of cyanobacteria. Upon nitrogen starvation, the iPGAM is inhibited by the PII-interacting regulator PirC, released from its interaction partner PII due to elevated 2-oxoglutarate levels. In-silico analysis of 338 different iPGAMs revealed a deep-rooted distinctive evolution of iPGAMs in cyanobacteria. Remarkably, cyanobacterial iPGAMs possess a unique loop structure and an extended C-terminus. Our analysis suggests that iPGAM forms a complex with three individual PirC monomers. Complex affinity is affected by the unique loop and the C-terminal structural elements. A C-terminal truncated enzyme showed loss of control by PirC and two-fold increased enzymatic activity compared to the iPGAM-WT. By contrast, deleting the loop structure drastically reduced the activity of this variant. By replacing the WT iPGAM in Synechocystis with different iPGAM variants, in which these structural elements were deleted, it became apparent that deletion of the C-terminal element showed a similar overproduction of polyhydroxybutyrate as deletion of the iPGAM-regulator PirC. However, in contrast to the latter, these strains showed higher over-all biomass accumulation, making them a better chassis for a production strain for PHB or other valuable substances than the PirC-deficient mutant. These findings significantly contribute to our understanding of the metabolic pathways in cyanobacteria and open up new avenues for further research in this field, inspiring future investigations and discoveries. The primordial cyanobacteria were responsible for developing oxygenic photo-synthesis early in evolution. Through endosymbiosis, they further evolved into the chloroplasts found in the plant kingdom. Many metabolic pathways within chloroplasts originated from cyanobacteria. However, differences emerged during their long separate evolution, providing insights into the endosymbiotic process. In the metabolic pathways involving fixed CO2, the co-factor-independent phosphoglycerate mutase (iPGAM) plays a crucial role by directing the first CO2 fixation product, 3-phosphoglycerate, towards critical anabolic path-ways. Our findings reveal a distinct evolution of iPGAM within oxygenic photo-synthetic organisms. We have identified two specific segments in cyanobacterial iPGAMs that tightly control the cellular carbon/nitrogen state through a specific protein interactor (PiC). This understanding of iPGAM has allowed us to engineer cyanobacterial strains with altered carbon fluxes. Since cyanobacteria can directly convert CO2 into valuable products, our results demonstrate a novel approach for developing a chassis for biotechnical use.

We propose a method-a quantum time mirror (QTM)-for simulating a partial time-reversal of the free-space motion of a non-relativistic quantum wave packet. The method is based on a short-time spatially homogeneous perturbation to the wave packet dynamics, achieved by adding a nonlinear time-dependent term to the underlying Schrödinger equation. Numerical calculations, supporting our analytical considerations, demonstrate the effectiveness of the proposed QTM for generating a time-reversed echo image of initially localized matter-wave packets in one and two spatial dimensions. We also discuss possible experimental realizations of the proposed QTM.

We propose a method -- a quantum time mirror (QTM) -- for simulating a partial time-reversal of the free-space motion of a nonrelativistic quantum wave packet. The method is based on a short-time spatially-homogeneous perturbation to the wave packet dynamics, achieved by adding a nonlinear time-dependent term to the underlying Schr\"odinger equation. Numerical calculations, supporting our analytical considerations, demonstrate the effectiveness of the proposed QTM for generating a time-reversed echo image of initially localized matter-wave packets in one and two spatial dimensions. We also discuss possible experimental realizations of the proposed QTM.

Both metaphysical and practical considerations related to time inversion have intrigued scientists for generations. Physicists have strived to devise and implement time-inversion protocols, in particular different forms of \"time mirrors\" for classical waves. Here we propose an instantaneous time mirror for quantum systems, i.e., a controlled time discontinuity generating wave function echoes with high fidelities. This concept exploits coherent particle-hole oscillations in a Dirac spectrum in order to achieve population reversal, and can be implemented in systems such as (real or artificial) graphene.