Biosynthesis of steroidal intermediates using Mycobacteria: a review / 生物工程学报

Steroids are a class of medicines with important physiological and pharmacological effects. In pharmaceutical industry, steroidal intermediates are mainly prepared through Mycobacteria transformation, and then modified chemically or enzymatically into advanced steroidal compounds. Compared with the "diosgenin-dienolone" route, Mycobacteria transformation has the advantages of abundant raw materials, cost-effective, short reaction route, high yield and environmental friendliness. Based on genomics and metabolomics, the key enzymes in the phytosterol degradation pathway of Mycobacteria and their catalytic mechanisms are further revealed, which makes it possible for Mycobacteria to be used as chassis cells. This review summarizes the progress in the discovery of steroid-converting enzymes from different species, the modification of Mycobacteria genes and the overexpression of heterologous genes, and the optimization and modification of Mycobacteria as chassis cells.

Advances in the production of chemicals by organelle compartmentalization in Saccharomyces cerevisiae / 生物工程学报

As a generally-recognized-as-safe microorganism, Saccharomyces cerevisiae is a widely studied chassis cell for the production of high-value or bulk chemicals in the field of synthetic biology. In recent years, a large number of synthesis pathways of chemicals have been established and optimized in S. cerevisiae by various metabolic engineering strategies, and the production of some chemicals have shown the potential of commercialization. As a eukaryote, S. cerevisiae has a complete inner membrane system and complex organelle compartments, and these compartments generally have higher concentrations of the precursor substrates (such as acetyl-CoA in mitochondria), or have sufficient enzymes, cofactors and energy which are required for the synthesis of some chemicals. These features may provide a more suitable physical and chemical environment for the biosynthesis of the targeted chemicals. However, the structural features of different organelles hinder the synthesis of specific chemicals. In order to ameliorate the efficiency of product biosynthesis, researchers have carried out a number of targeted modifications to the organelles grounded on an in-depth analysis of the characteristics of different organelles and the suitability of the production of target chemicals biosynthesis pathway to the organelles. In this review, the reconstruction and optimization of the biosynthesis pathways for production of chemicals by organelle mitochondria, peroxisome, golgi apparatus, endoplasmic reticulum, lipid droplets and vacuole compartmentalization in S. cerevisiae are reviewed in-depth. Current difficulties, challenges and future perspectives are highlighted.

Advances in abscisic acid biosynthesis / 生物工程学报

Abscisic acid, a plant hormone that inhibits growth, is a key factor in balancing plant endogenous hormones and regulating growth and metabolism. Abscisic acid can improve the drought resistance and salt tolerance of crops, reduce fruit browning, reduce the incidence rate of malaria and stimulate insulin secretion, so it has a broad application potential in agriculture and medicine. Compared with traditional plant extraction and chemical synthesis, abscisic acid synthesis by microorganisms is an economic and sustainable route. At present, a lot of progress has been made in the synthesis of abscisic acid by natural microorganisms such as Botrytis cinerea and Cercospora rosea, while the research on the synthesis of abscisic acid by engineered microorganisms is rarely reported. Saccharomyces cerevisiae, Yarrowia lipolytica and Escherichia coli are common hosts for heterologous synthesis of natural products due to their advantages of clear genetic background, easy operation and friendliness for industrial production. Therefore, the heterologous synthesis of abscisic acid by microorganisms is a more promising production method. The author reviews the research on the heterologous synthesis of abscisic acid by microorganisms from five aspects: selection of chassis cells, screening and expression enhancement of key enzymes, regulation of cofactors, enhancement of precursor supply and promotion of abscisic acid efflux. Finally, the future development direction of this field is prospected.

Advances of polymer-monomer production by cyanobacterial cell factory / 生物工程学报

Cyanobacteria is one of the promising microbial chassis in synthetic biology, which serves as a typical host for light-driven production. With the gradual depletion of fossil resources and intensification of global warming, the research on cyanobacterial cell factory using CO2 as carbon resource is ushering in a new wave. For a long time, research focus on cyanobacterial cell factory has mainly been the production of energy products, such as liquid fuels and hydrogen. One of the critical bottlenecks occurring in cyanobacterial cell factory is the poor economic performance, which is mainly caused by the inherent inefficiency of cyanobacteria. The problem is particularly prominent for these extremely cost-sensitive energy products. As an indispensable basis for modern industry, polymer monomers belong to the bulk chemicals with high added value. Therefore, increasing attention has been focused on polymer monomers which are superior in overcoming the economic barrier in commercialization of cyanobacterial cell factories. Here, we systematically review the progress on the production of polymer monomers using cyanobacteria, including the strategies for improving production, and the related technologies for the application of this important microbial cell factory. Finally, we summarize several issues in cyanobacterial synthetic biology and proposed future developing trends in this field.

Progress and perspective on development of non-model industrial bacteria as chassis cells for biochemical production in the synthetic biology era / 生物工程学报

The development and implement of microbial chassis cells can provide excellent cell factories for diverse industrial applications, which help achieve the goal of environmental protection and sustainable bioeconomy. The synthetic biology strategy of Design-Build-Test-Learn (DBTL) plays a crucial role on rational and/or semi-rational construction or modification of chassis cells to achieve the goals of "Building to Understand" and "Building for Applications". In this review, we briefly comment on the technical development of the DBTL cycle and the research progress of a few model microorganisms. We mainly focuse on non-model bacterial cell factories with potential industrial applications, which possess unique physiological and biochemical characteristics, capabilities of utilizing one-carbon compounds or of producing platform compounds efficiently. We also propose strategies for the efficient and effective construction and application of synthetic microbial cell factories securely in the synthetic biology era, which are to discover and integrate the advantages of model and non-model industrial microorganisms, to develop and deploy intelligent automated equipment for cost-effective high-throughput screening and characterization of chassis cells as well as big-data platforms for storing, retrieving, analyzing, simulating, integrating, and visualizing omics datasets at both molecular and phenotypic levels, so that we can build both high-quality digital cell models and optimized chassis cells to guide the rational design and construction of microbial cell factories for diverse industrial applications.

Advances in stress tolerance mechanisms and synthetic biology for the industrial robustness of Corynebacterium glutamicum / 生物工程学报

As a model industrial host and microorganism with the generally regarded as safe (GRAS) status, Corynebacterium glutamicum not only produces amino acids on a large scale in the fermentation industry, but also has the potential to produce various new products. C. glutamicum usually encounters various stresses in the process of producing compounds, which severely affect cell viability and production performance. The development of synthetic biology provides new technical means for improving the robustness of C. glutamicum. In this review, we discuss the tolerance mechanisms of C. glutamicum to various stresses in the fermentation process. At the same time, we highlight new synthetic biology strategies for boosting C. glutamicum robustness, including discovering new stress-resistant elements, modifying transcription factors, and using adaptive evolution strategies to mine stress-resistant functional modules. Finally, prospects of improving the robustness of engineered C. glutamicum strains ware provided, with an emphasis on biosensor, screening and design of transcription factors, and utilizing the multiple regulatory elements.

Construction of a highly efficient synthetic lycopene engineered Saccharomyces cerevisiae / 生物工程学报

Lycopene, as a high value-added terpene compound, has been widely concerned by researchers at home and abroad. Firstly, the ability of lycopene synthesis of Saccharomyces cerevisiae model strains S288c and YPH499 was analyzed and compared. The results showed that YPH499 was more suitable for lycopene synthesis as yeast chassis. Subsequently, the effects of constitutive promoters GPDpr, TEF1pr and inducible promoters GAL1pr, GAL10pr on Lycopene synthesis were compared. The results showed that when GPDpr and TEF1pr were used as promoters of crtE, crtB and crtI in lycopene synthesis pathway, the production of lycopene was 15.31 mg/L after 60 h fermentation in shaking flask. When GAL1pr and GAL10pr were used as promoters, the production was 123.89 mg/L, which was 8.09 times higher. In addition, the methylvaleric acid (MVA) pathway was further modified to overexpress the key enzyme gene of N-terminal truncation, tHMG1 (3-hydroxy-3-methylglutaryl coenzyme A reductase). The lycopene production was 265.68 mg/L, and the yield per cell was 72.79 mg/g. The Saccharomyces cerevisiae strain designed and constructed in this study can express lycopene in high yield per cell, thus could be used in the industrial production of lycopene after further construction and optimization.

Construction of seamless genome editing system for fast-growing Vibrio natriegens / 生物工程学报

Recently, fast-growing Vibrio natriegens, as the great potential chassis, has shown a wide application in synthetic biology. Genome editing is an indispensable tool for genetic modification in synthetic biology. However, genome editing tools with high efficiency and fidelity are still to be developed for V. natriegens synthetic biology. To deal with this problem, the physiological characteristics of 6 V. natriegens strains were evaluated, and CICC 10908 strain with fast and stable growth was selected as the host strain for genome editing study. Then, the natural transformation system of V. natriegens was established and optimized. The efficiencies of optimized natural transformation that integrates antibiotic resistance marker cat-sacB or Kan(R) onto the chromosome of V. natriegens could reach 4×10⁻⁵ and 4×10⁻⁴, respectively. Based on the optimized natural transformation, a double-selection cassette was used to achieve seamless genome editing with high efficiency and fidelity. The positive rates of four different types of genetic manipulation, including gene deletion, complementation, insertion and substitution, were 93.8%, 100%, 95.7% and 100%, respectively. Finally, transformation and elimination of the recombinant plasmid could be easily achieved in V. natriegens. This work provides a seamless genome editing system with high efficiency and fidelity for V. natriegens synthetic biology.

Genomics approaches decode natural products synthesis / 生物工程学报

Novel natural products have always been the most important sources for discovery of new drugs. Since the end of the 20th century, advances in genomics technology have contributed to decode and analyze numerous genomes, revealing remarkable potential for production of new natural products in organisms. However, this potential is hampered by laboratory culture conditions. Therefore, the integration of all these new advances is necessary to unveil these treasures, addressing the rise in resistance to antibiotics. In this review, we discuss the strategies of genome mining, inducing the expression of silent biosynthetic gene clusters and construction of biological chassis.

Nursing of patients with stoma chassis leakage and causes analysis / 现代临床护理

Objective To investigate the cause of stoma chassis leakage (SCL) and summarize the nursing experience. Method The clinical data of 20 patients with SCL were retrospectively analyzed on the colostomy outpatients. Result The causes of SCL in those stoma patients included insufficient skills of stoma care, peridermal depression, poorly fitting stoma chassis, weight gain, poor positioning and retraction of the stoma. Conclusion Such nursing measures as enhanced awareness of SCL causes, careful reselection of a suitable stoma chassis, addition of ointment patches or heightening of the convex chassis can be effective in prevention of SCL leakage, enhancement of patient′s confidence and improvement of their quality of life.