Microbial carbon fixation represents a crucial negative emission technology for achieving the carbon peaking and carbon neutrality goals and serves as a primary source of raw materials for the next generation of biomanufacturing. However, naturally occurring carbon fixation pathways generally suffer from limitations such as slow carboxylation rates, harsh reaction conditions, and difficulties in heterologous reconstruction. In recent years, scientists have designed and constructed several synthetic carbon fixation pathways, establishing a foundation for developing efficient biological carbon fixation systems. This article systematically reviews the naturally discovered carbon fixation pathways and summarizes the key advances in the modification of natural pathways and the design of synthetic carbon fixation pathways empowered by synthetic biology. Furthermore, it summarizes recent progress in energy supply strategies for microbial carbon fixation. Finally, we discuss the bottlenecks and optimization strategies, providing scientific insights and perspectives for the development of efficient carbon fixation systems for practical applications.

The valorization of one-carbon (C1) gases such as CO, CO₂,_{and syngas (a mixture with H2) represents a promising route for responding to global climate change and establishing a sustainable circular economy. Acetogens, a group of obligate anaerobic microorganisms capable of efficiently assimilating C1 gases via the Wood-Ljungdahl pathway (WLP), offer a natural conversion route with high carbon atom economy for biological carbon fixation. This paper provides a systematic review of the latest advancements in C1 gases conversion by acetogens, spanning from fundamental research to industrial applications. We first elucidate the metabolic coupling between acetogens and C1 gases, delving into the biochemical basis for utilizing C1 sources. In terms of fundamental research, we highlight recent breakthroughs in energy conservation mechanisms and metabolic flux regulation. Then, we summarize how the development of advanced genetic manipulation systems, exemplified by CRISPR-Cas tools, has enhanced the titers of natural products such as acetate and ethanol and accelerated the transformation of acetogens into chassis cells for the production of high-value chemicals such as butanol and 3-hydroxybutyrate. Finally, this review analyzes the engineering challenges, including gas-liquid mass transfer limitations and energy supply bottlenecks, associated with the scale-up of gas fermentation while offering a perspective on the broad application prospects of acetogens in achieving carbon peaking and carbon neutrality goals and advancing next-generation green biomanufacturing.}

Methane, the second most abundant greenhouse gas after carbon dioxide, represents a critical target in climate change mitigation efforts. Owing to their unique ability to utilize methane, methanotrophs have received substantial attention in greenhouse gas mitigation and low-carbon biomanufacturing, making the understanding of their metabolic pathways and ecological functions a vibrant research area worldwide. In recent years, the rapid development of high-throughput omics technologies and the integration of multi-omics analysis have accelerated a paradigm shift in methanotroph research, enabling the deep evolution from the identification of single gene functions to the elucidation of systemic metabolic processes. This review systematically summarizes the progress in the application of genomics, transcriptomics, proteomics, and metabolomics in methanotroph research, highlighting the decisive roles of omics technologies in elucidating carbon assimilation flux distribution in aerobic methanotrophs, uncovering novel metabolic pathways of anaerobic methane oxidation, and deciphering cross-domain interactions and electron transfer mechanisms within microbial consortia. To address current bottlenecks in the industrial application of methanotrophs, including poorly understood environmental adaptation mechanisms, imbalanced metabolic flux in chassis cells, and limited gas-liquid mass transfer efficiency, this review proposes a shift from descriptive omics toward function-driven and precision-intervention research. By integrating single-cell multi-omics and artificial intelligence-enhanced modeling, future studies may construct efficient artificial methanotrophic cell factories and synthetic microbial communities, thereby providing innovative biological solutions for achieving the global carbon neutrality goal.

Against the backdrop of global climate change and energy security, developing efficient carbon dioxide utilization technologies has become a strategic core for achieving the “dual carbon” goals. Photosynthetic biomanufacturing, which uses photoautotrophic organisms as chassis to directly convert solar energy and carbon dioxide into biofuels and bio-based chemicals, enables simultaneous carbon fixation, emission reduction, and green synthesis. It stands as a pivotal technology for advancing carbon neutrality. Cyanobacteriota, with their efficient photosynthetic capacity, well-defined genetic background, and mature operational systems, have emerged as highly promising photosynthetic cell chassis. To advance their large-scale application, there is an urgent need to develop highly efficient cyanobacteria-driven photosynthetic carbon fixation cell factories. This review systematically summarizes recent research progress from two dimensions: enhancing photosynthetic carbon fixation efficiency and achieving precise carbon flux direction toward target products. In terms of photosynthetic carbon fixation, this review mainly focuses on strategies such as endogenous pathway optimization, external material-enabled enhancement, and chassis exploration and reshaping for constructing efficient and stable photosynthetic carbon fixation systems. Regarding carbon flux direction and product synthesis, this review analyzes the implementation pathways for precise carbon resource orientation and efficient synthesis of complex products, centered on single-strain metabolic engineering and multi-strain modular collaboration. Finally, this review outlines future development directions in this field. The review aims to provide theoretical references and technical pathways for the systematic construction of cyanobacteria-driven photosynthetic carbon fixation cell factories, thereby contributing to the achievement of carbon neutrality.

Against the backdrop of global climate change and resource scarcity, biological carbon fixation technologies driven by clean energy have emerged and attracted widespread attention. Hydrogen (H₂), as a renewable and clean energy carrier with abundant sources and easy production, can serve as both the energy and the electron donor for microbial carbon fixation. This review summarizes the metabolic foundations of hydrogen-driven carbon-fixing chassis cells represented by hydrogen-oxidizing bacteria (HOB), and highlights recent advances in several representative strains. Furthermore, engineering strategies for HOB are discussed, including the development of enabling technologies, the reconstruction and reinforcement of metabolic pathways, and microbe-material coupling approaches. Additionally, this paper analyzes the current challenges facing HOB and proposes the future perspectives. Despite existing technical bottlenecks, with the advancement of metabolic engineering and systems biology, HOB are expected to play an important role in carbon recycling and sustainable biomanufacturing.

One-carbon (C1) compounds, including CO₂, methanol, and formate, are regarded as promising feedstocks for sustainable biomanufacturing. Diverse natural carbon fixation pathways, such as the Calvin-Benson-Bassham (CBB) cycle, the reductive tricarboxylic acid (rTCA) cycle, and the Wood-Ljungdahl pathway (WLP), have been extensively investigated. However, their inherent limitations in energy efficiency and carbon fixation rate have constrained the development of efficient C1 biomanufacturing systems, thereby motivating the rapid development of artificial C1 assimilation pathways. In recent years, a variety of artificial C1 assimilation pathways have been designed for the utilization of CO₂, methanol, and formate, and their functions have been validated in both in vitro and in vivo systems, providing new approaches for the construction of efficient C1-based biomanufacturing platforms. This review systematically summarizes recent advances in artificial C1 assimilation pathways using CO₂, methanol, and formate as substrates, with a particular focus on pathway design, the engineering and optimization of key enzymatic components, and the construction and application of artificial pathways in microbial chassis. In addition, this paper discusses the future perspectives on artificial carbon fixation, providing guidance and insights for the development of efficient and sustainable C1 biomanufacturing systems.

With the proposal of carbon neutrality goals, microbial conversion of CO₂ into high-value chemicals, fuels, and biomaterials has emerged as an important strategy for clean energy and green manufacturing. Although single-strain carbon fixation systems have ideal controllability, they still face limitations, including heavy metabolic burden, limited carbon fixation flux, insufficient energy and reducing power supply, low yield, and challenges in industrial-scale application. In contrast, synthetic microbial consortia enable rational metabolic division of labor among functionally distinct microorganisms, thereby alleviating metabolic burden, enhancing carbon fixation efficiency, and expanding product diversity. These advantages further improve product value and promote the industrial application of CO₂ fixation. This review summarizes recent advances in the application of synthetic microbial consortia for CO₂ conversion, including photosynthesis-driven systems, hydrogen-driven systems, and potentially safer chemoautotrophic systems classified according to carbon fixation modules. In addition, this paper discusses the design principles for constructing carbon cycling systems based on synthetic microbial consortia, together with current challenges and future perspectives, providing theoretical and practical guidance for the development of efficient and stable artificial carbon cycling systems.

Under the strategic goal of carbon neutrality, enhancing the carbon atom utilization efficiency of microbial cell factories has emerged as a core scientific challenge and imperative technical demand for advanced green biomanufacturing. Conventional fermentation processes possess mature industrial applicability, yet they are constrained by inevitable carbon losses via oxidative decarboxylation in central carbon metabolism. Such inherent limitation hinders fundamental improvement in substrate carbon conversion efficiency and restricts the green and high-quality advancement of the biomanufacturing industry. Multi-carbon source co-fermentation enables the rational integration of carbohydrate substrates with C1 feedstocks (formic acid, methanol, and syngas) and C2 feedstocks (acetic acid), constructing a mixotrophic metabolic system featured with carbon skeleton complementation, energy supply synergy, and intracellular redox balance. This strategy offers an innovative technological paradigm to break the theoretical carbon yield bottleneck of conventional bioprocesses. This paper systematically reviews the advances in multi-carbon source co-fermentation driven by synthetic biology. We emphatically elaborate on metabolic pathway reconstruction and regulatory mechanisms of typical co-fermentation systems consisting of organic substrates, C1/C2 compounds, and industrial waste gas. The state-of-the-art applications in synthetic fuels, bio-based materials, and high-value natural product biosynthesis are summarized. Furthermore, this paper discusses the industrial potential of this technology in integrating carbon capture and utilization and high-value biomanufacturing, aiming to provide a theoretical basis and strategic references for the innovation of efficient biomanufacturing towards carbon neutrality.

Achieving carbon peaking and carbon neutrality is a major strategic priority for China. Third-generation biomanufacturing, which uses one-carbon (C1) compounds such as methanol, carbon dioxide, and formic acid as feedstocks for bioconversion, has attracted increasing research interest. Among these C1 feedstocks, methanol is considered a promising substrate for biomanufacturing because of its ease of storage and transportation, high degree of reduction, and potential for large-scale production via CO₂ hydrogenation. This review systematically compares natural and synthetic methanol assimilation pathways and elucidates their advantages. It further summarizes methanol metabolism in natural methylotrophic microorganisms and pathway design strategies for constructing synthetic methylotrophs, while also discussing the major challenges associated with methanol utilization. On this basis, this paper reviews recent progress in the methanol-based biosynthesis of various high-value chemicals and discusses the bottlenecks and corresponding engineering strategies for methanol bioconversion. This review provides a theoretical foundation for methanol-driven green biomanufacturing.

Methanol is regarded as an ideal non-food feedstock for biomanufacturing due to its abundant supply and low cost. Methylorubrum extorquens AM1 has become a prominent model chassis strain in this field, owing to its natural ability to utilize methanol. This review summarizes recent advances in methanol-based biomanufacturing viaM. extorquens AM1 cell factories. First, the methanol metabolic pathways and compatible synthetic biology tools available for this strain are discussed. Subsequently, we highlight recent achievements in the metabolic engineering of this strain for the production of recombinant proteins, biodegradable materials, organic acids, and terpenoids. Building on these findings, we outline the metabolic engineering strategies that have been employed to enhance the biosynthetic capacity of engineered strains. Finally, we discuss the current limitations in constructing and applying M. extorquens AM1 cell factories and propose future research directions. This review will serve as a valuable reference for the development of M. extorquens cell factories and for the industrial production of methanol-derived products.