Terpenoids are a class of natural products with important physiological functions and significant biological activities that are widely found in nature and have a wide range of applications in the food, medical, and daily chemical industries. In the biosynthetic pathway of terpenoids, terpene synthases often determine the type and novelty of the terpene carbon skeleton, and tailoring enzymes, such as cytochrome P450 enzymes, can carry out a variety of post-modifications, ultimately resulting in terpenoids with a rich diversity of structures and functions. In recent years, with the development of genome-sequencing technology and synthetic biology, a large number of terpene biosynthetic enzymes of plant and microbial origin have been characterized, which, excitingly, include non-canonical terpene synthases that can also catalyze the generation of unique cyclized skeletons. Meanwhile, the use of combinatorial biosynthetic strategies has led to the creation of many novel and unnatural terpenoids, further enriching the kingdom of terpenoids. Here, we review the recent advances in non-canonical terpene cyclases and combinatorial biosynthetic pathways over the past five years, with a view to shedding light on the discovery and biosynthesis of novel terpenes in the future. Firstly, the newly discovered novel enzymes with terpene cyclization functions are reviewed, containing a new subclass of type Ⅰ terpene synthases, non-squalene triterpene synthases, UbiA-type terpene cyclases, cytochrome P450 oxygenases, methyltransferases, vanadium-dependent haloperoxidases, and haloacid dehalogenase, along with their sequences, functions, and possible cyclization mechanisms, which can contribute to our understanding of terpenoid biosynthetic enzymes and the discovery of novel terpenoids. This review then describes the combinatorial biosynthesis of non-canonical terpenoids. By combining terpene synthases with methyltransferases or natural/artificial cytochrome P450 oxygenases, a series of unnatural terpenoids containing non-canonical C₁₁ and C₁₆ backbones, or with unusual structural modifications, were produced. This could inspire the structural innovation studies of terpenoids in the future. The discovery of novel enzymes and the construction of novel combinatorial biosynthetic pathways will further broaden the structural diversity and chemical space of terpenoids, which is expected to provide more potential novel terpenoids for clinical drug development.

Glycosylation modifications, extensively present on the surfaces of eukaryotic proteins as a type of post-translational modification, hold significant physiological and pathological implications. The microscopic heterogeneity of natural glycoproteins has led to the emergence of the chemical synthesis of homogeneous glycoproteins with defined structures as a crucial frontier in exploring the structure-function relationships of glycosylation modifications. With the flourishing development of protein synthesis and glycoengineering technologies, various protein ligation and polysaccharide synthesis strategies have been developed, enabling the preparation of glycoproteins containing hundreds of amino acid residues. The development of glycoprotein synthesis strategies primarily revolves around chemical and enzymatic approaches for glycosidic bond formation, leading to effective synthesis schemes such as Lansbury’s aspartic acid acylation, chemical strategies based on glycosyl amino acid building blocks, and glycan remodeling strategies using endoglycosidases and glycosyltransferases. This review will discuss the chemical and enzymatic construction of glycosidic bonds, examining existing strategies for the total synthesis of glycoproteins and semi-synthetic approaches that combine with biological expression methods. It will introduce these strategies’ achievements in synthesizing complex homogeneous glycoproteins with different types of glycosylation modifications, such as those with multiple complex N-glycosylation modifications like HSV gD and those containing long hydrophobic segments like IL-2. Additionally, this review will highlight breakthroughs in understanding the structure-function relationships of glycosylation modifications in various physiological processes through these synthetic complex glycoproteins, including the relationship between glycan chain length and immunogenicity in antigenic glycoproteins, and the mechanisms by which O-GlcNAc regulates synaptic function in neurons. Finally, it will summarize the progress made in glycosidic bond construction, purification strategies, and protein solubility, and point out that further optimization of selectivity and synthetic yield remains a pressing issue in the field of glycoprotein synthesis. The wide application of glycoprotein synthesis technology in developing immunotherapies and understanding the molecular mechanisms of various diseases expands the development directions of synthetic science in the field of life and health, from understanding principles to developing products.

DNA-Encoded Library (DEL) technology, as an emerging means of small molecule drug screening, has become an important and indispensable technology platform for new drug discovery and development. The technology incorporates many advantages from combinatorial chemistry, molecular biology, and chemical bioinformatics, which greatly improve the efficiency of compound library synthesis and screening. Meanwhile, driven by the development of nucleic acid-compatible chemical reactions and high-throughput sequencing technology, DEL technology has made remarkable progress and gradually become a fast, economical, and efficient high-throughput screening platform, and has been more and more widely used in seedling compounds screening by universities, research institutes, and large pharmaceutical companies. The success of a DEL screening relies heavily on the chemical space and structural diversity of the compound libraries, both of which are directly affected by the number of chemical reactions compatible with nucleic acids. Therefore, developing the on-DNA chemical reactions to continuously enrich the chemical toolbox for DEL synthesis and thus enhance the structural diversity and drug potential of the molecules in the libraries has been the focus in this field. In recent years, the number of on-DNA chemical reactions has increased significantly, greatly broadening the scope of chemical reactions available for DEL construction. Meanwhile, a series of novel reaction methods, such as photocatalysis, electrocatalysis, and biosynthesis, have also emerged in the application of on-DNA chemical reactions and further expanded the field that on-DNA chemical reactions can reach. In this paper, we systematically review the metal-catalyzed on-DNA chemical reactions in recent years, including C(sp²)—C(sp²) bond-formation reactions, C(sp³)—C(sp³) bond-formation reactions, C(sp²)—C(sp³) bond-formation reactions, and C(sp²)—X bond-formation reactions; the synthesis of on-DNA privileged heterocycles with single-ring, fused-ring, and spirocyclic rings by using target-oriented synthetic and diversity-oriented synthetic strategies; the research progress of photocatalytic and enzyme-catalyzed on-DNA chemical reactions. However, the current developments in on-DNA reactions also have limitations, such as compatibility with nucleic acids and substrate suitability. In the future, it is important to exploit more robust on-DNA reactions that can proceed under mild conditions, new types of on-DNA reactions, and the combination of high-throughput screening and computer-assisted on-DNA reactions.

Advances in science and technology are creating huge benefits and value for society. The digitalization of the world has brought great changes to human being’s daily life. Meanwhile, the increasing degree of digitalization has led to an unprecedented explosion of data, resulting in increasingly severe information storage challenges. According to the current developing trend, the global data volume is expected to reach 175 zettabytes by 2025. With the rapid growth of global data volume and the exponential growth of total data, the existing storage methods will no longer be able to meet the storage needs brought by the digitalization of the world and then there is an urgent need to develop information storage methods with better storage performance, higher storage efficiency and more durable storage media. Nature has offered a powerful solution by using DNA molecules as carriers of information, where genetic information has been transferred stably more than a million years. DNA storage has many advantages over traditional storage media, including high storage density, potentially low maintenance costs, and ease of synthesis and chemical modification, which make it an ideal alternative for information storage. The current process of storing data in DNA includes six main steps: encoding, writing, preservation, retrieval, reading, and decoding. Among them, the writing of data is the basic for realizing the storage of data in DNA, concluding writing data in DNA sequence and in DNA structure. In this review, we first introduce strategies for in vivo data writing in DNA storage systems, which primarily involve writing data into DNA sequences and DNA structures. This is followed by an overview of the development of in vivo writing techniques in DNA storage systems. Finally, we discuss the challenges faced by DNA storage systems in terms of high writing costs and slow writing speeds, and prospects for large-scale synthesis of high-purity DNA and improved biocatalysts.

In order to achieve carbon neutrality and green economy, people use biorefinery technology to transform and utilize CO₂. Microbial electrosynthesis (MES) is an emerging technology that converts CO₂ into chemicals by electrically driven biocatalysts. Currently, the low efficiency of microbial carbon sequestration, an incomplete understanding of electron transfer mechanisms, low synthesis rate, and poor applicability of reactor components have been the limiting factors for the large-scale application of MES. In this paper, the mechanisms of electron supply in the MES system, including through electrodes and electron donors such as H₂, formic acid, CO, and other molecules, are systematically reviewed based on how cathodic microorganisms obtain electrons. It is an effective method to improve electron transport efficiency by modifying conductive nanowires of electroactive microorganisms and optimizing the expression of microbially associated hydrogenase, formate dehydrogenase and CO dehydrogenase using synthetic biology techniques. Additionally, cathode modification aimed at improving electron transfer rates between microbes and electrodes, enhancing the biocompatibility, and providing more reducing power can facilitate the generation of value-added products. In addition to enhancing the electron transfer efficiency of the cathode, the construction of a reactor with high efficiency of gas-liquid-solid mass transfer and electron transfer, the reduction of anode potential for water electrolysis, and the regulation of microbial activity are also important strategies to enhance MES performance. In the future, it is necessary to further elucidate the mechanism of microbial electron transport and strengthen the performance of MES by means of synthetic biological communities, and by designing a more efficient electrode interface that balances electron transfer rate, substrate mass transfer and biocompatibility. In terms of the scaling-up of reaction devices, electron transfer and gas mass transfer can be improved through the combination of various methods, and integrating product separation processes can promote the further development of the technology and provide new ideas for the realization of the “Carbon Peak and Carbon Neutrality” goal.

Organic acids, as important platform chemicals, have been widely used in food, pharmaceutical, chemical industries and agriculture. Currently, microbial production of organic acids relies primarily on sugars as feedstocks, which may suffer from the competition with food and arable lands. One carbon (C₁) molecules such as CO, CO₂, methane, methanol and formic acid are widespread and inexpensive, which are considered as ideal feedstocks for future bio-manufacturing. Bioconversion of C₁ feedstocks toward the production of organic acids helps mitigate greenhouse effect and realize carbon neutrality. Therefore C₁ sources have been regarded as raw materials of third generation biorefinery, and natural C₁ utilizing microbes attracted increasing attention. Although some microorganisms have native biosynthetic pathway of organic acids, the production efficiency is usually lower than expected. This review summarizes the recent progress on the biosynthesis of organic acids (3-hydroxypropionic acid, lactic acid and succinic acid) from C₁ feedstocks using synthetic biology methods. First, the native C₁ utilizing pathways are summarized, including CO₂, CO, methane, methanol and formic acid. Then the metabolic engineering strategies to improve organic acids production were systematically reviewed, including the optimization of rate-limiting enzymes expression, enhancement of the supply of precursor and cofactor, cofactor engineering, and inhibition of the product degradation. In addition, the challenges, solutions, and prospects of C₁ bioconversion to organic acids are also discussed, and coupling chemical catalysis and biological transformation may provide a promising industrial route for organic acids production. In particular, methanol is an ideal C₁ feedstock with many advantages like convenient storage and transportation, high liquid-to-liquid mass transfer efficiency, and it can also be massively produced from CO₂ by “liquid sunshine” technology. Therefore constructing high efficient methanol cell factory may enable organic acids production from CO₂, a carbon neutral production manner. This review may provide a guidance for C₁ biorefinery and industrial bioproduction of organic acids.

The extensive consumption of fossil oil and the rapid accumulation of greenhouse gas emissions have caused long-term changes in the global climate and environment, sparking widespread interest in society for CO₂ bioconversion technologies as a means to address energy transition and climate change. As a new-generation biorefinery platform based on synthetic biology, the synthetic phototrophic community comprises closely cooperating phototrophic and heterotrophic microorganisms. This community is capable of efficiently converting light energy directly into biomass and a variety of chemicals through mutualistic metabolic division of labor among community members. Synthetic phototrophic community is one of the potential ways to achieve sustainable carbon-negative biomanufacturing, and has attracted widespread attention attributed to its advantages in applicability and robustness. In recent years, with the rapid development of systems biology and synthetic biotechnology, a variety of research efforts have been applied to the design and optimization of synthetic phototrophic communities, achieving stable progress and promoting the understanding of phototrophic community production. In this review, we briefly introduced an overview of the advances and current status of synthetic phototrophic community, including mutualistic mechanisms related to element, energy, and information flow. Subsequently, the unique advantages of phototrophic community were outlined. Meanwhile, recent systems biology approaches of phototrophic community were summarized, such as integrative analysis of multi-omics data, genome-scale metabolic modelling, flux balance analysis and community performance predictive algorithms. We also focused on the design and optimization strategies, such as chassis upgrading, immobilization/compartmentalization techniques, and enhanced internal multilayer regulation of synthetic phototrophic community, as well as the progress of their applications in various fields. Furthermore, we analyzed and discussed the constraints and challenges for the further deployment of synthetic phototrophic community on a larger scale, ranging from photosynthetic carbon production rate, intermediate organic matter selection, external predator invasion, to light distribution under high density cultivation. Finally, the future research strategies and engineering directions of synthetic phototrophic community encompassing semiconductor biohybrids, fine regulation of interspecies interaction and multi-omics community model construction were proposed. We conclude by providing a perspective on the future application scenarios of synthetic phototrophic communities in biochemistry, biomedicine, bioremediation and bioagriculture.

Synthetic biology offers boundless possibilities and revolutionary changes to material fields. One remarkable outcome of interdisciplinary integration of synthetic biology and material science is the development of environmentally friendly polyhydroxyalkanoates (PHAs), which serve as ideal alternatives to petroleum-based plastics. PHAs are a family of linear biopolyesters synthesized by various microorganisms as their intracellular storage materials for energy and carbon sources. With at least 150 various monomers, PHAs exhibit diverse structures, material properties, and applications, collectively known as “PHAomics”. When reprograming microbial genomes via synthetic biology and metabolic engineering, in combination with the feeding of special precursors, tailor-made PHAs with defined structures and varied properties can be synthesized. PHAs has been extensively studied in both academia and industry in the last few decades, leading to the commercialization of some PHAs. Next generation industrial biotechnology (NGIB) based on halophilic Halomonas spp. as chassis has been developed to overcome the limitations of current industrial biotechnology. NGIB offers a long lasting, open and continuous, energy and freshwater-saving bioprocess using low-cost mixed substrates and allows morphology engineering for simplified downstream processing. NGIB facilitates low-cost production of various PHAs in large scale. This review introduces PHAomics and summarizes the diverse properties of PHAs produced via NGIB. It primarily focuses on the composition, structure, and material properties of PHAs, as well as their extensive applications in biodegradable plastics, medical implants, medicine, drug delivery carriers, energy sources, and potential smart materials. Additionally, it covers the strategies and tools for strain engineering and their achievements in the tailor-made biosynthesis of PHA using reprogrammed Pseudomonas spp. and Halomonas spp. Finally, this review discusses strategies on how to further reduce the production cost and improve material properties of PHAs. This review summarizes the progresses on the low-cost customized synthesis of PHA biomaterials by synthetic biology, demonstrating the integration of biology and chemistry.

The early development of the human embryo includes three important stages: ①the pre-implantation stage from the zygote to the late blastocyst; ②the peri-implantation stage from late blastocyst to pre-gastrulation embryo; and ③the most mysterious post-gastrulation stage from gastrulation to early organogenesis. The latter two stages are collectively referred to as the early post-implantation developmental stage. During pregnancy, infertility (implantation failure or miscarriage) and birth defects of the fetus are largely due to abnormalities in human early postimplantation development. Human early postimplantation embryo, due to its small size and location in the mother’s uterus, is difficult to observe and study. Therefore, the embryonic development process at this stage has been in a black box state for a long time. In recent years, with the emergence of single-cell omics technology and extended in vitro culture system of human blastocysts, as well as the rapid development in the fields of embryonic and extraembryonic stem cells, organoids and embryoids, the mystery of the human early postimplantation development is gradually being lifted. In order to help understand the mysteries of early human embryonic development, this review primarily introduces the lineage diversification, key developmental events and known developmental principles during early human embryogenesis; summarizes recent progress in the research on human embryonic and extraembryonic stem cells (including totipotent stem cells, embryonic stem cells, trophoblast stem cells, primitive endoderm stem cells and extraembryonic mesoderm cells); presents the effects of cell communication, lineage interaction, signal gradient, adhesion molecules, biomechanics, and extracellular matrix on cell sorting, migration rearrangement and self-organization in embryoids and organoids; reviews the current research status of human early post-implantation embryogenesis, stem cell-based embryo models and organoids; and finally proposes the prospects and possible solutions to the problems and challenges existing in the research of human early post-implantation development using stem cell-derived embryo models or organoids.

Human early embryonic development is critical for a healthy fetus birth. However, the specific molecular regulatory mechanisms of lineage development, cell fate decisions and embryonic patterning are still shrouded in mystery. Our knowledge about early human embryogenesis has been greatly improved with the recent progress in in vitro culture conditions for human blastocysts and the advancements in omics technology. However, ethical and technological challenges continue to pose obstacles in these studies. With the rapid development of human pluripotent stem cells, they can be coaxed to form embryo-like structures that mimic early human embryonic development in vitro, termed “embryo models”. Interest in human embryonic development has been reinvigorated with the continuing advances in this area. Human embryo models can be divided into two categories "non-integrated embryo models" and “integrated embryo models” according to the different cellular components they possessed. Integrated embryo models represent the embryo-like structures containing both embryonic and extra-embryonic cell types, including blastoids, human extra-embryoids (hEEs), E-assembloids, stem-cell-derived synthetic whole embryo models (SEMs), peri-gastruloids, bilaminiods and heX-embryoids. While non-integrated embryo models sometimes lack the extra-embryonic tissues, including embryoid bodies (EBs), gastruloids, micro-patterned colonies, post-implantation amniotic sac embryoids (PASE). Besides, non-human primate cynomolgus monkey models have greatly expanded our knowledge towards human developmental biology. In this review, we summarized recent human stem cell-based non-integrated and integrated embryo models, and pointed out the technical challenges remained with proposed future directions. These findings lay an important foundation for understanding early human embryonic development, promoting research into human stem cells and their application, as well as preventing and treating early pregnancy loss or congenital diseases. On the other hand, embryo-like structures derived from human stem cell-based integrated embryo models, although do not fully, almost recapitulate the key events and structural organization of the in vivo counterparts, thus eliciting a serious compact on traditional ethical and practical concerns. In the future, non-integrated human embryo models and non-human primate models, which pose fewer ethical challenges, may provide a straightforward way to study the human early embryonic development.