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Quick Search     Advanced Search   2024 Vol.  33 No.  5 Published: 2024-10-28 1 2024 Vol. 33 (5): 1-2 [Abstract] ( 23 ) [HTML 1KB] [PDF 726KB] ( 136 ) 385 LIU Jiezheng, LIU Min, ZHAO Guang, XIAN Mo Progress in Artificial Microbiol Transformation of One-carbon Compounds Abstract: One-carbon compounds (C1), including methane, formate, methanol, carbon monoxide and carbon dioxide, have been considered to be the ideal raw materials for the third generation of biorefining, due to their abundance and low cost. Microbes that can naturally utilize C1 have been widely studied with low carbon conversion rate and industrial difficulty. The transformation and heterologous expression of C1 utilization pathway described as synthetic one-carbon bio-utilization system can help overcome the obstacles of non-model organisms, which will reduce the dependence on traditional fossil fuels. This review briefly summarized the types, sources and natural pathways of C1. We mainly focused on the progress of C1 utilization in model organisms and emphasized the importance of synthetic C1 bio-utilization systems for improving carbon conversion rate and developing new application. In addition, we discussed the challenges and perspectives of the biotransformation of C1. This paper provided a reference for the construction of synthetic C1 utilization microorganisms and improving the efficiency of heterologous C1 metabolic pathway, which is of great significance for realizing green and efficient development of sustainable resources. Key words: one-carbon compounds; biotransformation; carbon dioxide; synthetic one-carbon bio-utilization system; carbon conversion rate （Acta Laser Biology Sinica, 2024, 33(5): 385-399） 2024 Vol. 33 (5): 385-399 [Abstract] ( 34 ) [HTML 1KB] [PDF 4217KB] ( 199 ) 400 FANG Jing, RANG Jie, XIA Liqiu Synthetic Biology Applications for Enhancing the Efficient Production of Spinosad in Saccharopolyspora spinosa Abstract: Spinosad, a secondary metabolite produced by Saccharopolyspora spinosa, has garnered significant attention globally due to its environmentally friendly and broad-spectrum insecticidal properties. The unique structure of spinosad and its analog, butenyl-spinosyn, underlies their distinctive mode of action. To meet the demands of large-scale industrial production, research teams worldwide have been striving to enhance the yields of spinosad and butenyl-spinosyn, with remarkable achievements accomplished through the application of synthetic biology techniques. This review summarizes the research conducted domestically and internationally on the regulation of metabolic pathways at various levels using synthetic biology techniques to improve the production of spinosad and butenyl-spinosyn. It focuses on four key aspects: chassis cell modification and optimization, multi-omics analysis, genetic circuit modification, and metabolic network regulation. Furthermore, it provides an outlook on future research directions in this field, and provides a new research strategy to promote the biosynthesis of spinosad by synthetic biology technology. Key words: spinosad; Saccharopolyspora spinosa; synthetic biology; gene editing; metabolic pathway regulation （Acta Laser Biology Sinica, 2024, 33(5): 400-407） 2024 Vol. 33 (5): 400-407 [Abstract] ( 41 ) [HTML 1KB] [PDF 1617KB] ( 174 ) 408 CUI Ying, SONG Kai, HE Yawen L-methionine Biosynthesis and Regulatory Mechanisms in Bacteria Abstract: L-methionine is an essential amino acid with a wide range of applications in food, feed, cosmetics and pharmaceuticals. Currently, L-methionine is the only essential amino acid that cannot be industrially produced by microbial fermentation. In recent years, the use of metabolic engineering to enhance the yield of L-methionine has received widespread attention from researchers at home and abroad. In this paper, the biosynthetic pathway and regulatory mechanism of L-methionine in Corynebacterium glutamicum and Escherichia coli are analysed; three key steps (i.e., acylation, sulfurylation, and methylation) in the biosynthesis of methionine from hyper-serine are highlighted, and a further outlook on the biosynthesis of L-methionine is proposed with a view to providing guidance for the industrial production of L-methionine. Key words: L-methionine; biosynthesis; regulatory mechanism （Acta Laser Biology Sinica, 2024, 33(5): 408-417） 2024 Vol. 33 (5): 408-417 [Abstract] ( 40 ) [HTML 1KB] [PDF 2676KB] ( 185 ) 418 LIN Yaqian, SHUAI Feifei, WANG Fang Application of Synthetic Microorganisms in Water Pollution Prevention and Control Abstract: Water pollution is one of the most serious environment problems that affect people′s life quality and sustainable economic development worldwide. The degradation capability of microorganisms endows them advantages in environmental pollutants elimination and water environments repairment, with the utilization of diverse bioremediation technologies. However, due to the complex situation of water pollution environment, traditional microbiological treatment methodology is restricted and limited. With the engineering principle Design-Build-Test-Learn in synthetic biology, the designed and modified synthetic microorganisms with efficient and broad-spectrum degradation ability for pollutants has attracted the researchers′ attention. The synthetic microorganisms could not only meet the wastewater treatment requirements within complex pollutants, but also could achieve the green, efficient circular treatment goal. Therefore, design and construction of synthetic microorganisms for water pollution control has becoming one of the future development and application direction of synthetic biology. Herein, we introduced the current situation of water pollution, the treatment technologies classically utilized, and the synthetic microorganisms′ application in water pollution control. This review focuses on different types of microbial chassis in synthetic biology and the merits of application of synthetic microorganisms in water pollution control. This review will facilitate the performance optimization of pollutant degradation and wastewater treatment via synthetic biology and provide reference for the efficient treatment of wastewater environment. Key words: synthetic microorganisms; microbial chassis; water pollution; bioremediation technology; environmental governance （Acta Laser Biology Sinica, 2024, 33(5): 418-431） 2024 Vol. 33 (5): 418-431 [Abstract] ( 48 ) [HTML 1KB] [PDF 2755KB] ( 167 ) 432 CHEN Meiying, XIE Yuheng, TANG Miaomiao, XI Xuedong Research Progress on Methylase Modification in the Synthesis of Natural Products#br# Abstract: Methyltransferases (MTs) are a class of enzymes that are ubiquitous in biological organisms, usually using S-adenosylmethionine as a methyl donor to catalyze the methylation reaction of the substrate. The heterologous expression of MTs in microorganisms has made great progress in realizing the biosynthesis of some important natural products. MTs can be used in microorganisms to synthesize important natural products such as phenylpropanoids, fragrance compounds, hormones and antibiotics. MTs have also been widely used in many fields such as medicine, chemical industry and energy, showing great application value and broad application prospects. In this review, we summarize the classification, function and application of natural product methyltransferases, in order to provide theoretical guidance for the efficient artificial biosynthesis of highly active non-ribosomal peptide synthetase (RXPs) peptides. Key words: methyltransferases; S-adenosylmethionine; methylation; natural products; biosynthesis （Acta Laser Biology Sinica, 2024, 33(5): 432-438） 2024 Vol. 33 (5): 432-438 [Abstract] ( 45 ) [HTML 1KB] [PDF 2054KB] ( 147 ) 439 SHEN Zhuoling, WAN Muyang, DENG Lei A RHD1 Variant Lacking Deubiquitination Function Retains Specific Binding Activity to Linear Ubiquitin Chain Abstract: The special connection approaches of linear ubiquitin chain have been elucidated in recent years. Currently, the types of commercial linear ubiquitin chain specific antibodies are limited and their specificity still needs to be improved. RHD1 is a papain-like deubiquitinating enzyme that specifically recognizes and cleaves linear ubiquitin chains. The mutation at C13 in its enzymatic activity site results in the loss of deubiquitination function for the RHD1. Meanwhile, the RHD1 with this mutation still exhibits low binding activity towards linear ubiquitin chain. On the basis of this mutant, we optimized the mutations at interaction interface between RHD1 and Di-Ub (M1-linked) in order to enhance the specific binding activity of RHD1. In this study, we utilized the structural model of RHD1 and Di-Ub (M1-linked) complex to analyze the physical and chemical properties of all amino acids at the interaction interface and subsequently designed a RHD1 mutant library. The site-mutations in RHD1 variant polypeptides were generated by using overlap extension PCR method, and all recombinant variant proteins were expressed by using E. coli BL21 (DE3) strain. By employing the enzyme-linked immune sorbent assay (ELISA ) method for screening, we successfully identified a RHD1 variant harboring C13A and A92T mutations, which showed significantly increased affinity to Tetra-Ub (M1-linked). This RHD1 variant holds great potential as a valuable tool for studying linear ubiquitin chains, and exhibits promising application in clinical diagnostics and the development of related therapeutic agents. Key words: RHD1 deubiquitinating enzymes; Tetra-Ub (M1-linked); site-specific mutation; prokaryotic expression; ELISA （Acta Laser Biology Sinica, 2024, 33(5): 439-449） 2024 Vol. 33 (5): 439-449 [Abstract] ( 53 ) [HTML 1KB] [PDF 3880KB] ( 178 ) 450 CHEN Wangqiong, ZHU Yan, ZHAO Qin, LIU Xirong, XIA Liqiu, RANG Jie Important Functional Genes Mining Regulating Spinosyn Biosynthesis and Its Action Mechanism Abstract: Synthetic biology can fully promote the efficient bio-manufacturing of target natural products in the aspects of efficient chassis cell construction, active natural product mining and metabolic pathway optimization. However, in the study of synthetic biology of Saccharopolyspora spinosa, due to the limited understanding of the important functional genes regulating spinosyn biosynthesis and their action mechanisms, it is difficult to construct efficient cell factories to significantly increase the spinosyn production through the “design-build-test-learn” strategy. In order to solve this problem, an excellent chassis strain CW-12 with high spinosyn production, fast growth rate and strong extracellular glucose uptake ability was obtained by artificial mutagenesis, and comparative proteomic analysis was performed. The results showed that the enhancement of metabolic pathways related to intracellular carbon metabolism, fatty acid metabolism, amino acid biosynthesis, and TCA cycle was an important reason for the improvement of spinosyn biosynthesis of mutant CW-12, and 668 up-regulated proteins were screened. Subsequently, we selected 3-hydroxyacyl-CoA dehydrogenase (SS_2202) and acetyl-CoA acetyltransferase (SS_2203) to analyze their action mechanisms, confirming that their overexpression can effectively promote the spinosyn biosynthesis. This study has certain guiding significance on how to explore the important functional genes regulating spinosyn biosynthesis and the analysis of their action mechanisms, and lays an important foundation for the subsequent construction of spinosyn efficient biosynthesis cell factories through synthetic biology strategies. Key word: Saccharopolyspora spinosa; spinosyn; synthetic biology; comparative proteomic analysis; gene editing （Acta Laser Biology Sinica, 2024, 33(5): 450-460） 2024 Vol. 33 (5): 450-460 [Abstract] ( 52 ) [HTML 1KB] [PDF 3822KB] ( 165 ) 461 YU Jia, ZHANG Liang, LIU Tianbo, CAI Hailin, YI Ke, QIU Liting, WANG Yunsheng, LIU Qingshu, CHEN Wu The Two-component System YvrG/YvrH Regulates the Biosynthesis of Edeine Abstract: Edeine, synthesized by Brevibacillus brevis X23, is a linear peptide antibiotic not encoded by ribosomal RNA, renowned for its extensive range of antibacterial effectiveness. Transcriptome sequencing revealed an inverse expression pattern between the two-component regulatory factors yvrG/yvrH and the edeine biosynthesis gene cluster (ede BGC), suggesting the involvement of YvrH/YvrG in the transcriptional regulation of ede BGC. This study utilized Red/ET homologous recombination technology to construct X23 mutant strains that yvrG/yvrH lacks, complemented strains, and overexpressing strains, and investigated the impact of YvrG/YvrH on edeine biosynthesis through phenotype analysis and quantitative real-time PCR (RT-qPCR) techniques. The results revealed that knocking out yvrG/yvrH significantly reduced the antibacterial activity of the strains, the expression of ede BGC decreased, the synthesis of edeine was inhibited, and the yield of edeine was reduced by 46.75%. In summary, YvrG/YvrH act as positive regulatory factors in the edeine biosynthesis. This study provides gene regulatory elements for the metabolic engineering of edeine. Key words: Brevibacillus brevis; edeine; two-component regulatory factors; bacteriostatic activity; biosynthesis （Acta Laser Biology Sinica, 2024, 33(5): 461-469） 2024 Vol. 33 (5): 461-469 [Abstract] ( 35 ) [HTML 1KB] [PDF 5449KB] ( 143 ) 470 XIA Lun, ZHU Yan, WANG Shanrui, LIU Xirong, XIA Liqiu, RANG Jie Effect of GluA on Growth Development and Butenyl-spinosyn Biosynthesis in Saccharopolyspora pogona Abstract: Glutamate transporter GluA, is a class of ATP-binding proteins that can affect intracellular carbon metabolism, nitrogen metabolism and secondary metabolism by regulating the active transport of glutamate, belonging to the ABC transport system. To study the effects of GluA on the strain growth development and butenyl-spinosyn biosynthesis in Saccharopolyspora pogona, gluA-1 overexpression and deletion strains were constructed by genetic engineering technology. Phenotypic analysis showed that the butenyl-spinosyn production increased by 4.2 times, the bacterial growth logarithmic phase was prolonged, the maximum bacterial density and acetyl-CoA content significantly increased, whereas the mycelium morphology was not significantly changed in overexpressed strain S. pogona::gluA-1. The butenyl-spinosyn production decreased by 37%, the strain growth was inhibited obviously, the acetyl-CoA content significantly decreased, and the mycelium was shortened and thickened, branching was reduced in deletion strain S. pogona-ΔgluA-1. These results indicated that GluA could promote the growth development and the butenyl-spinosyn biosynthesis in S. pogona, which laid an important foundation for the study of GluA's role in the secondary metabolism of Streptomyces. Key word: Saccharopolyspora pogona; butenyl-spinosyn; glutamate transporter protein; biology synthesis; gene editing （Acta Laser Biology Sinica, 2024, 33(5): 470-480） 2024 Vol. 33 (5): 470-480 [Abstract] ( 54 ) [HTML 1KB] [PDF 3994KB] ( 166 )

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