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Journal of Central South University 2009年第1期16卷 53-60页

作者：夏金兰孟浩王润民张成桂熊晶聂珍媛邱冠周 Key Laboratory of Biometallurgy of Ministry of Education Central South University School of Minerals Processing and Bioengineering Central South University

In order to solve the citrus peel resource waste problem protopectinase-overproducing strain CD-01 for pectin production and minimize the drawbacks of chemical extraction of pectin, a was isolated from a pit soil dumped with perished orange in Changde City, Hunan Province of China. The strain CD-01 had the same morphology and 28S rRNA gene sequence （FJ184995） as that of Aspergillus niger （ATCC 64028）. It was thus identified and named as Aspergillus niger CD-01. The fermentation condition was optimized based on L9（34） orthogonal experimental design and the variances analyses. The results show that the optimal condition for producing pectin is as follows： time 36 h, temperature 35 ℃, pH 5, and urea as the nitrogen source. Under this condition, the pectin yield can reach up to 24.5%. This shows a great potential of Aspergillus niger CD-01 in pectin extraction from citrus.

关键词： Aspergilllus niger pectin production protopectinase citrus peel fermentation optimization

Engineering Komagataella phaffii to biosynthesize cordycepin from methanol which drives global metabolic alterations at the transcription level

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Synthetic and Systems Biotechnology 2023年第2期8卷 242-252页

作者： Huiping Tan Liang Wang Huiguo Wang Yanghao Cheng Xiang Li Huihui Wan Chenguang Liu Tian Liu Qian Li Department of Life and Health Dalian UniversityNo.10 Xuefu StreetDalian Economic and Technological Development ZoneDalian116622China School of Biological Engineering Dalian Polytechnic UniversityNo.1 Qinggongyuan RoadGanjingzi DistrictDalian116034China Analytical Instrumentation Centre Dalian University of TechnologyNo.2 Linggong RoadDalian116024China State Key Laboratory of Microbial Metabolism Joint International Research Laboratory of Metabolic&Developmental ScienceSchool of Life Sciences and BiotechnologyShanghai Jiao Tong University800 Dongchuan RoadMinhang DistrictShanghai200240China School of Bioengineering Dalian University of TechnologyNo.2 Linggong RoadDalian116024China

Cordycepin has the potential to be an alternative to the disputed herbicide ***,current laborious and time-consuming production strategies at low yields based on Cordyceps militaris lead to extremely high cost and restrict its application in the field of *** this study,Komagataella phaffii(*** pastoris)was engineered to biosynthesize cordycepin from methanol,which could be converted from CO_(2).Combined with fermentation optimization,cordycepin content in broth reached as high as 2.68±0.04 g/L within 168 h,around 15.95 mg/(L⋅h)in ***,a deaminated product of cordycepin was identified at neutral or weakly alkaline starting pH during *** analysis found the yeast producing cordycepin was experiencing severe inhibition in methanol assimilation and peroxisome biogenesis,responsible for delayed growth and decreased carbon flux to pentose phosphate pathway(PPP)which led to lack of precursor *** acid interconversion and disruption in RNA metabolism were also due to accumu-lation of *** study provided a unique platform for the manufacture of cordycepin based on the emerging non-conventional yeast and gave practical strategies for further optimization of the microbial cell factory.

关键词： Cordycepin Komagataella phaffii fermentation optimization Transcriptome

Coupled strategy based on regulator manipulation and medium optimization empowers the biosynthetic overproduction of lincomycin

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Synthetic and Systems Biotechnology 2024年第1期9卷 134-143页

作者： Xinlu Cai Wanlian Xu Yang Zheng Sendi Wu Rundong Zhao Nian Wang Yaqian Tang Meilan Ke Qianjin Kang Linquan Bai Buchang Zhang Hang Wu School of Life Sciences Institute of Physical Science and Information TechnologyAnhui UniversityHefei230601China State Key Laboratory of Microbial Metabolism Shanghai Jiao Tong UniversityShanghai200240China

The biosynthesis of bioactive secondary metabolites,specifically antibiotics,is of great scientific and economic *** control of antibiotic production typically involves different processes and molecular *** numerous efforts to improve antibiotic yields,joint engineering strategies for combining genetic manipulation with fermentation optimization remain *** A(Lin-A),a lincosamide antibiotic,is industrially fermented by Streptomyces ***,the leucine-responsive regulatory protein(Lrp)-type regulator SLCG_4846 was confirmed to directly inhibit the lincomycin biosynthesis,whereas indirectly controlled the transcription of SLCG_2919,the first reported repressor in *** of SLCG_4846 in the high-yield *** LA219X(LA219XΔ4846)increases the Lin-A production and deletion of SLCG_2919 in LA219XΔ4846 exhibits superimposed yield *** the effect of the double deletion on cellular primary metabolism of ***,Plackett-Burman design,steepest ascent and response surface methodologies were utilized and employed to optimize the seed medium of this double mutant in shake flask,and Lin-A yield using optimal seed medium was significantly increased over the *** strategies were performed in a 15-L *** maximal yield of Lin-A in LA219XΔ4846-2919 reached 6.56 g/L at 216 h,55.1%higher than that in LA219X at the parental cultivation(4.23 g/L).This study not only showcases the potential of this strategy to boost lincomycin production,but also could empower the development of high-performance actinomycetes for other antibiotics.

关键词： Streptomyces Transcription factor Genetic engineering fermentation optimization Lincomycin

Characterization of putative mannoprotein in Kluyveromyces lactis for lactase production

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Synthetic and Systems Biotechnology 2023年第1期8卷 168-175页

作者： Xiuru Shen Lingtong Liao Guoqiang Zhang Jingwen Zhou Jianghua Li Guocheng Du Science Center for Future Foods Jiangnan UniversityWuxi214122China National Engineering Research Center for Cereal fermentation and Food Biomanufacturing Jiangnan University1800 Lihu RoadWuxiJiangsu214122China School of Biotechnology and Key Laboratory of Industrial Biotechnology Ministry of EducationJiangnan University1800 Lihu RoadWuxiJiangsu214122China

Lactase is a member of theβ-galactosidase family of enzymes that can hydrolyze lactose into galactose and ***,extracellular lactase production was still restricted to the process of cell *** this study,lactase-producing Kluyveromyces lactis JNXR-2101 was obtained using a rapid and sensitive method based on the fluorescent substrate 4-methylumbelliferyl-β-*** purified enzyme was identified as a neutral lactase with an optimum pH of *** facilitate extracellular production of lactase,a putative mannoprotein KLLA0_E01057g of *** was knocked *** could effectively promote cell wall degradation and lactase production after lyticase treatment,which showed potential on other extracellular enzyme *** optimizing the fermentation conditions,the lactase yield from mannoprotein-deficient *** JNXR-2101ΔE01057g reached 159.62 U/mL in a 5-L fed-batch bioreactor.

关键词： Kluyveromyces lactis Lactase Screening Enzymatic properties Mannoprotein fermentation optimization Secretion

Optimizing production ofasperolide A,a potential anti-tumor tetranorditerpenoid originally produced by the algal-derived endophytic fungus Aspergillus wentii EN-48

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Chinese Journal of Oceanology and Limnology 2017年第3期35卷 658-663页

作者：徐蕊李晓明徐刚明王斌贵 Laboratory of Marine Biology and Biotechnology Qingdao National Laboratory for Marine Science and TechnologyKey Laboratory of Experimental Marine BiologyInstitute of OceanologyChinese Academy of SciencesQingdao 266071China University of Chinese Academy of Sciences Beijing 100049China

The marine algal-derived endophytic fungus Aspergillus wentii EN-48 produces the potential anti-tumor agent asperolide A, a tetranorlabdane diterpenoid active against lung cancer. However, the fermentation yield of asperolide A was very low and only produced in static cultures. Static fermentation conditions of A. wentii EN-48 were optimized employing response surface methodology to enhance the production of asperolide A. The optimized conditions resulted in a 13.9-fold yield enhancement, which matched the predicted value, and the optimized conditions were successfully used in scale-up fermentation for the production of asperolide A. Exogenous addition of plant hormones （especially 10 pmol/L methyl jasmonate） stimulated asperolide A production. To our knowledge, this is first optimized production of an asperolide by a marine-derived fungus. The optimization is effective and valuable to supply material for further anti-tumor mechanism studies and preclinical evaluation of asperolide A and other norditerpenoids.

关键词： Aspergillus wentii asperolide A anti-tumor agent fermentation optimization

fermentation Performance and Characterization of Cold-Adapted Lipase Produced with Pseudomonas Lip35

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Agricultural Sciences in China 2009年第8期8卷 956-962页

作者： YU Hong-wei HAN Jun LI Ning QIE Xiao-sha JIAYing-min Institute of Food Science and Technology Agricultural University of Hebei Baoding 071001 PR.China

Strain of Pseudomonas Lip35 producing lipase was isolated in a refrigerator. Lipase production and characterization of this strain were investigated under different conditions. The Pseudomonas was cultivated in shaking flasks in a fermentation medium in various nutritional and physical environments. Lipase production has been influenced by the presence of yeast-extract, soybean powder, NaCI, and Tween-80. Maximum lipase productivity was obtained when the physical environment of the fermentation medium was optimal for 67 h. The production of lipase reached 58.9 U·mL^-1. The lipase of Pseudomonas Lip35 can be considered to be inducible, but the inducer had little influence on the production of lipase. The lipase was characterized and showed high lipolytic activity from pH 7.5-8.0. The optimum temperature was observed at 20℃ and the thermal inactivation of lipase was obvious at 60℃. The lipase activity was inhibited by K＋, stimulated by Ca^2＋, and thermostability decreased in the presence of Ca^2＋, therefore the lipase was Ca^2＋ -dependent cold-adapted enzyme.

关键词： cold-adapted lipase fermentation optimization lipase characterization Pseudomonas Lip35

Combinatorial mutagenesis of Bacillus amyloliquefaciens for efficient production of protease

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Systems Microbiology and Biomanufacturing 2023年第3期3卷 457-468页

作者： Kuo Zhao Haoyu Liu Wei Song Jing Wu Cong Gao Liang Guo Xiulai Chen State Key Laboratory of Food Science and Technology Jiangnan University1800 Lihu RoadWuxi 214122China International Joint Laboratory On Food Safety Jiangnan UniversityWuxi 214122China School of Life Sciences and Health Engineering Jiangnan UniversityWuxi 214122China

As an important industrial enzyme,protease is widely used in feed,food and other *** present,the insufficient protease activity obtained from microorganisms cannot meet the purpose of industrial *** this study,Bacillus amyloliquefaciens with high protease production was screened from animal feces by plate transparent circle *** improve the production of protease,atmospheric room temperature plasma(ARTP)mutagenesis was used in the first round,protease activity reached 315.0 U/***,to enhance production of protease,^(60)Co-γ irradiation was used for combined mutagenesis,leading to protease activity of *** FMME ZK003 up to 355.0 U/***,to realize the efficient production of protease,after optimization of fermentation conditions,protease activity was increased to 456.9 U/***,protease activity of *** FMME ZK003 reached 823.0 U/mL in a 5 L *** results indicate that *** can efficiently produce protease,which provides a good foundation for the industrial production of protease.

关键词： Bacillus amyloliquefaciens Protease production Atmospheric room temperature plasma ^(60)Co-γirradiation fermentation optimization

Genomics-guided discovery of a new and significantly better source of anticancer natural drug FK228

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Synthetic and Systems Biotechnology 2018年第4期3卷 268-274页

作者： Xiangyang Liu Feng Xie Leah B.Doughty Qi Wang Lixin Zhang Xueting Liu Yi-Qiang Cheng State Key Laboratory of Bioreactor Engineering East China University of Science and TechnologyShanghai200237PR China UNT System College of Pharmacy University of North Texas Health Science CenterFort WorthTX76107USA CAS Key Laboratory of Pathogenic Microbiology and Immunology Institute of MicrobiologyChinese Academy of SciencesBeijing100101PR China Department of Biological Sciences University of Wisconsin-MilwaukeeMilwaukeeWI53201USA

FK228 is an FDA-approved anticancer drug naturally produced by Chromobacterium violaceum No.968 up to 19 mg/L in a pilot industry-scale batch *** we report a genomics-guided discovery of Burkholderia thailandensis MSMB43 as a new and significantly better source of *** genome of *** MSMB43 was found to contain a functional biosynthetic gene cluster highly homologous to that of FK228 in *** No.968,and the bacterium indeed produces authentic *** simple fermentation in shaking flasks in a preferred M8 medium,*** MSMB43 produced FK228 up to 67.7 mg/L;by fedbatch fermentation in a 20-L fermentor in M8 medium,*** MSMB43 produced FK228 up to 115.9 mg/L,which is 95 fold higher than that of *** No.968 under the same laboratory fermentation ***-PCR analysis indicated that the high FK228 yield of *** MSMB43 was due to high expression of biosynthetic genes,represented by Bth_depA,during the fermentation *** genetic manipulation resulted in a recombinant strain,*** MSMB43/pBMTL3-tdpR,which harbors a broad host-range vector expressing the thailandepsin biosynthetic pathway regulatory gene *** engineered strain produced up to 168.5 mg/L of FK228 in fed-batch fermentation in a 20-L fermentor in M8 ***,the wild-type *** MSMB43 or its engineered derivative could potentially be a good starting point for an industrial process to improve FK228 production for its expanding use in therapy.

关键词： Burkholderia thailandensis MSMB43 fermentation optimization FK228 Genome mining Natural product Productivity

Constitutive expression of codon optimized Trichoderma reesei TrCel5A in Pichia pastoris using GAP promoter

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Systems Microbiology and Biomanufacturing 2022年第3期2卷 498-506页

作者： Yun Hu Renhui Bai Shaohua Dou Zhimeng Wu Ali Abdulkhani Mohammad Ali Asadollahi Abd El-Fatah Abomohra Fubao Sun Key Laboratory of Carbohydrate and Biotechnology and Key Laboratory of Industrial BiotechnologyMinistry of EducationSchool of BiotechnologyJiangnan UniversityWuxi 214122China School of Life Science and Biotechnology Dalian UniversityDalian 116622China Department of Wood and Paper Sciences and Technology Faculty of Natural ResourcesUniversity of Tehran31585-4314 KarajIran Department of Biotechnology Faculty of Biological Science and TechnologyUniversity of Isfahan81746-73441 IsfahanIran New Energy and Environmental Laboratory School of Architecture and Civil EngineeringChengdu UniversityChengdu 610106China

To address the deficient activity of TrCel5A in naturally secreted cellulase preparation,this study used the GAP promoter to induce constitutive expression of Trichoderma reesei TrCel5A in Pichia pastoris.A recombinant TrCel5A was screened out after gene optimization,synthesis,and *** biochemical and enzymatic properties of the new recombinant were *** a result,optimization of shake-flask fermentation of the recombinant was obtained at 28℃,2%inoculum volume,an initial pH of 6.0,as well as glycerol and Tween-80 additions of 30 g/L and 6 g/L,*** the above-optimized conditions,the recombinant produced 14.8 U/mL of the enzyme activity at 96 h of *** further enhance enzyme production,pilot-scale cultivation was evaluated using 5-L *** high-cell-density fermentation,the recombinant strain increased enzyme activity to 130.4 U/ml and protein content to 2.49 g/*** addition,the kinetic factors,including K_(m) and V_(max) values for TrCel5A,were detected to be 5.1 mg/mL and 265.9μmol/(***),***,TrCel5A was effectively expressed in *** under the GAP promoter,and it demonstrated its potential in commercially relevant enzyme hydrolysis of lignocellulosic biomass.

关键词： Trichoderma reesei endoglucanase(Cel5A) Pichia pastoris GAP promoter Constitutive expression fermentation optimization CMC enzyme activity

Statistical culture-based strategies to enhance chlamydospore production by Trichoderma harzianum SH2303 in liquid fermentation

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Journal of Zhejiang University-Science B(Biomedicine & Biotechnology) 2016年第8期17卷 619-627页

作者： Ya-qian LI Kai SONG Ya-chai LI Jie CHEN School of Agriculture and Biology Shanghai Jiao Tong University Shanghai 200240 China Key Laboratory of Urban Agriculture (South) Ministry of Agriculture Shanghai 200240 China Affiliated Hospital of Hebei University Baoding 071100 China

Trichoderma-based formulations are applied as commercial biocontrol agents for soil-borne plant path- ogens. Chlamydospores are active propagules in Trichoderma spp., but their production is currently limited due to a lack of optimal liquid fermentation technology. In this study, we explored response surface methodologies for opti- mizing fermentation technology in Trichoderma SH2303. Our initial studies, using the Plackett-Burman design, iden- tified cornmeal, glycerol, and initial pH levels as the most significant factors （P〈0.05） for enhancing the production of chlamydospores. Subsequently, we applied the Box-Behnken design to study the interactions between, and optimal levels of, a number of factors in chlamydospore production. These statistically predicted results indicated that the highest number of chlamydospores （3.6×108 spores/ml） would be obtained under the following condition： corn flour 62.86 g/L, glycerol 7.54 ml/L, pH 4.17, and 6-d incubation in liquid fermentation. We validated these predicted values via three repeated experiments using the optimal culture and achieved maximum chlamydospores of 4.5×108 spores/ml, which approximately a 8-fold increase in the number of chlamydospores produced by T. harzianurn SH2303 compared with that before optimization. These optimized values could help make chlamydospore production cost-efficient in the future development of novel biocontrol agents.

关键词： Trichoderma harzianum SH2303 Chlamydospore Plackett-Burman screening Box-Behnken design fermentation optimization