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A conservative distribution of tridomain NDP-heptose synthetases in actinobacteria

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Science China(Life Sciences) 2022年第5期65卷 1014-1023页

作者： Yue Tang Wei Tang Min Wang Zhilong Zhang Yihua Chen State key laboratory of microbial Resources and cas key laboratory of microbial physiological and Metabolic engineering Institute of MicrobiologyChinese Academy of SciencesBeijing 100101China University of Chinese Academy of Sciences Beijing 100049China

Heptoses are important structural components of Gram-negative bacterium cell wall and participate in bacterial colonization,infection,and immune *** knowledge of NDP-heptose originating from D-sedoheptulose 7-phosphate in Grampositive bacterium remains ***,in silico analysis suggested that the special tridomain NDP-heptose synthetases with isomerase,kinase,and nucleotidyltransferase activities are conservatively distributed in Actinobacteria class of Gram-positive *** characterization of the tridomain proteins from different strains showed that they are involved in ADPD-glycero-β-D-manno-heptose biosynthesis despite the unexpected discovery of kinase activities deficient in some *** presence of three types of NDP-heptose synthetases in Gram-positive bacterium suggests that it is also a rich source of heptoses and the heptose moieties may play important roles in *** work updates the understanding of NDP-heptose biosynthesis in Gram-positive bacterium and lays a solid foundation for further physiological function explorations.

关键词： tridomain NDP-heptose synthetase NDP-heptose biosynthesis widespread distribution Gram-positive bacterium

Traceless enzymatic protein synthesis without ligation sites constraint

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National Science Review 2022年第5期9卷 108-118页

作者： Ruifeng Li Marcel Schmidt Tong Zhu Xinyu Yang Jing Feng Yu'e Tian Yinglu Cui Timo Nuijens Bian Wu cas key laboratory of microbial physiological and Metabolic engineering State Key Laboratory of Microbial ResourcesInstitute of Microbiology Chinese Academy of Sciences University of Chinese Academy of Sciences Fresenius Kabi iPSUM I&D Center EnzyPep B.V.

Protein synthesis and semisynthesis offer immense promise for life sciences and have impacted pharmaceutical *** absence of a generally applicable method for traceless peptide conjugation with a flexible choice of junction sites remains a bottleneck for accessing many important synthetic targets,*** we introduce the PALME(protein activation and ligation with multiple enzymes) platform designed for sequence-unconstrained synthesis and modification of *** upstream activating modules accept and process easily accessible synthetic peptides and recombinant proteins,avoiding the challenges associated with preparation and manipulation of activated peptide ***,the downstream coupling module provides comprehensive solutions for sequential peptide condensation,cyclization and protein N/C-terminal or internal *** practical utility of this methodology is demonstrated by synthesizing a series of bioactive targets ranging from pharmaceutical ingredients to synthetically challenging *** modular PALME platform exhibits unprecedentedly broad accessibility for traceless protein synthesis and functionalization,and holds enormous potential to extend the scope of protein chemistry and synthetic biology.

关键词： protein synthesis traceless peptide ligation biocatalysis synthetic biology chemical biology

Turning waste gases into valuables

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Synthetic and Systems Biotechnology 2022年第3期7卷 847-848页

作者： Huawei Zhu Yin Li cas key laboratory of microbial physiological and Metabolic engineering State Key Laboratory of Microbial ResourcesInstitute of MicrobiologyChinese Academy of SciencesBeijing100101China

Waste off-gases from steel mills and other mining related industries often contain rich carbon monoxide(CO),carbon dioxide(CO_(2)),and sometimes hydrogen(H2).The biological conversion of waste gases into chemicals is an absolutely carbon-negative *** date,the only chemical that can be commercially produced from fermentation of waste gas is ethanol,an important two-carbon(C2)chemical generally used as a blending component of gasoline[1].This technology was developed by LanzaTech *** has been operating on a commercial scale since ***,another breakthrough in gas fermentation was reported by Liew et al.[2]from the same *** achieved pilot-scale production of acetone and isopropanol from steel mill off-gas by using an engineered acetogen Clostridium autoethanogenum.

关键词： steel gases breakthrough

α-Glucosidase Inhibitory Activities of Lutein and Zeaxanthin Purified from Green Alga Chlorella ellipsoidea

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Journal of Ocean University of China 2018年第4期17卷 983-989页

作者： QI Jia KIM Sang Moo cas key laboratory of microbial physiological and Metabolic engineering Institute of MicrobiologyChinese Academy of Sciences Department of Marine Food Science and Technology Gangneung-Wonju National University

α-Glucosidase inhibitors are used therapeutically to treat type-2 diabetes mellitus. Through a bioassay-guided fractionation technique, three carotenoids,(all-E)-lutein,(all-E)-zeaxanthin and(9-Z)-zeaxanthin, were purified from the green alga Chlorella ellipsoidea, in which(all-E)-lutein and(9-Z)-zeaxanthin had potent α-glucosidase inhibitory activity. IC_(50) values of(all-E)-lutein and(9-Z)-zeaxanthin were 70 and 53.5 μmol L^(-1) against Saccharomyces cerevisiae α-glucosidase, respectively, with non-competitive inhibition. In addition, IC_(50) values of(9-Z)-zeaxanthin against Bacillus stearothermophilus and rat-intestinal α-glucosidase were 805.1 and 671.2 μmol L^(-1), respectively. The K_i values of(all-E)-lutein and(9-Z)-zeaxanthin against S. cerevisiae α-glucosidase were 78.1 and 16.5 μmol L^(-1), respectively. Therefore, C. ellipsoidea carotenoids might be utilized as a novel candidate to prevent type-2 diabetes mellitus related disorders in food and medical industry.

关键词： type-2 diabetes mellitus α-glucosidase inhibitor carotenoids Chlorella ellipsoidea

Synthetic biology for CO2 fixation

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Science China(Life Sciences) 2016年第11期59卷 1106-1114页

作者： Fuyu Gong Zhen Cai Yin Li cas key laboratory of microbial physiological and Metabolic engineering Institute of MicrobiologyChinese Academy of SciencesBeijing 100101China University of the Chinese Academy of Sciences Beijing 100049China

Recycling of carbon dioxide(CO_2) into fuels and chemicals is a potential approach to reduce CO_2 emission and fossil-fuel consumption. Autotrophic microbes can utilize energy from light, hydrogen, or sulfur to assimilate atmospheric CO_2 into organic compounds at ambient temperature and pressure. This provides a feasible way for biological production of fuels and chemicals from CO_2 under normal conditions. Recently great progress has been made in this research area, and dozens of CO_2-derived fuels and chemicals have been reported to be synthesized by autotrophic microbes. This is accompanied by investigations into natural CO_2-fixation pathways and the rapid development of new technologies in synthetic biology. This review first summarizes the six natural CO_2-fixation pathways reported to date, followed by an overview of recent progress in the design and engineering of CO_2-fixation pathways as well as energy supply patterns using the concept and tools of synthetic biology. Finally, we will discuss future prospects in biological fixation of CO_2.

关键词： carbon dioxide fixation synthetic biology CO2-fixation pathway energy supply

Developing controllable hypermutable Clostridium cells through manipulating its methyl-directed mismatch repair system

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Protein & Cell 2013年第11期4卷 854-862页

作者： Guodong Luan Zhen Cai Fuyu Gong Hongjun Dong Zhao Lin Yanping Zhang Yin Li cas key laboratory of microbial physiological and Metabolic engineering Institute of MicrobiologyChinese Academy of SciencesBeijing 100101China University of Chinese Academy of Sciences Beijing 100049China

Development of controllable hypermutable cells can greatly benefit understanding and harnessing microbial ***,there have not been any similar sys-tems developed for Clostridium,an important bacterial *** we report a novel two-step strategy for de-veloping controllable hypermutable cells of Clostridium acetobutylicum,an important and representative indus-trial ***,the mutS/L operon essential for methyl-directed mismatch repair(MMR)activity was inactivated from the genome of *** to generate hy-permutable cells with over 250-fold increased mutation ***,a proofreading control system carrying an inducibly expressed mutS/L operon was *** hypermutable cells and the proofreading control system were integrated to form a controllable hypermut-able system SMBMutC,of which the mutation rates can be regulated by the concentration of anhydrotetracycline(aTc).Duplication of the miniPthl-tetR module of the proof-reading control system further signifi cantly expanded the regulatory space of the mutation rates,demonstrating hypermutable Clostridium cells with controllable mutation rates are *** developed *** strain SMBMutC2 showed higher survival capacities than the control strain facing butanol-stress,indicating greatly increased evolvability and adaptability of the controllable hypermutable cells under environmental challenges.

关键词： Clostridium acetobutylicum mutation rates hypermutable cells artifi cial control

Enzymatic clickable functionalization of peptides via computationally engineered peptide amidase

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Chinese Chemical Letters 2018年第7期29卷 1116-1118页

作者： Tong Zhu Lu Song Ruifeng Li Bian Wu cas key laboratory of microbial physiological and Metabolic engineering State Key Laboratory of Microbial ResourcesInstitute of MicrobiologyChinese Academy of SciencesBeijing 100101China University of Chinese Academy of Sciences Beijing 100101China State key laboratory of Transducer Technology Chinese Academy of SciencesBeijing 100101China

Directed peptides C-terminal modification enabled by the engineered biomolecular catalyst-peptide amidase 12 B has been achieved via computational protein engineering. The engineered enzyme exhibits great promising potential in the C-terminal modification of opioid peptides using prop-2-yn-1-amine(PYA) or prop-2-en-l-amine(PEA) as the nucleophile. A variety of opioid peptides could be readily functionalized at the C-terminal chain in high yield in a mild and selective manner. Notably, modified opioid peptides bearing alkynyl moiety could be further functionalized through well-established click reaction.

关键词： Peptide amidase Opioid peptides C-terminal modification Click reaction Functionalization

Development of an activity-directed selection system enabled significant improvement of the carboxylation efficiency of Rubisco

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Protein & Cell 2014年第7期5卷 552-562页

作者： Zhen Cai Guoxia Liu Junli Zhang Yin Li cas key laboratory of microbial physiological and Metabolic engineering Institute of Microbiology Chinese Academy of Sciences Beijing 100101 China Tianjin Institute of Industrial Biotechnology Chinese Academy of Sciences Tianjin 300308 China

Photosynthetic CO2 fixation is the ultimate source of organic carbon on earth and thus is essential for crop production and carbon sequestration, Ribulose-1,5-bis- phosphate carboxylase/oxygenase （Rubisco） catalyzes the first step of photosynthetic CO2 fixation. However, the extreme low carboxylation efficiency of Rubisco makes it the most attractive target for improving pho- tosynthetic efficiency. Extensive studies have focused on re-engineering a more efficient enzyme, but the effort has been impeded by the limited understanding of its structure-function relationships and the lack of an effi- cient selection system towards its activity. To address the unsuccessful molecular engineering of Rubisco, we developed an Escherichia coil-based activity-directed selection system which links the growth of host cell solely to the Rubisco activity therein. A Synechococcus sp. PCC7002 Rubisco mutant with E49V and D82G sub- stitutions in the small subunit was selected from a total of 15,000 mutants by one round of evolution. This mutant showed an 85% increase in specific carboxyla- tion activity and a 45% improvement in catalytic efficiency towards CO2. The small-subunit E49V mutation was speculated to influence holoenzyme catalysis through interaction with the large-subunit Q225. This interaction is conserved among various Rubisco from higher plants and Chlamydomonas reinhardtii. Knowledge of these might provide clues for engineering Rubisco from higher plants, with the potential of increasing the crop yield.

关键词： carboxylation efficiency CO2 fixation directed evolution Rubisco Synechococcus sp. PCC7002

Genetic biosensors for small-molecule products： Design and applications in high-throughput screening

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Frontiers of Chemical Science and engineering 2017年第1期11卷 15-26页

作者： Qingzhuo Wang Shuang-Yan Tang Sheng Yang Institute of Plant Physiology and Ecology Shanghai Institutes for Biological Sciences Shanghai 200025 China University of the Chinese Academy of Sciences Beijing 100049 China cas key laboratory of microbial physiological and metebolic engineering Institute of Microbiology Chinese Academy of Sciences Beijing 100101 China Jiangsu National Synergetic Innovation Center for Advanced Materials Nanjing Tech University Nanjing 211816 China

Overproduction of small-molecule chemicals using engineered microbial cells has greatly reduced the production cost and promoted environmental protection. Notably, the rapid and sensitive evaluation of the in vivo concentrations of the desired products greatly facilitates the optimization process of cell factories. For this purpose, many genetic components have been adapted into in vivo biosensors of small molecules, which couple the intracel- lular concentrations of small molecules to easily detectable readouts such as fluorescence, absorbance, and cell growth. Such biosensors allow a high-throughput screen- ing of the small-molecule products, and can be roughly classified as protein-based and RNA-based biosensors. This review summarizes the recent developments in the design and applications of biosensors for small-molecule products.

关键词： biosensor small molecule product transcrip-tion factor riboswitch high-throughput screening