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Yanwen Duan - One of the best experts on this subject based on the ideXlab platform.
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streptomycin induced ribosome engineering complemented with Fermentation Optimization for enhanced production of 10 membered enediynes tiancimycin a and tiancimycin d
Biotechnology and Bioengineering, 2019Co-Authors: Zhoukang Zhuang, Yanwen Duan, Rong Huang, Yong Huang, Chengzhou Jiang, Fan Zhang, Xiaohui Yan, Xiangcheng ZhuAbstract:Tiancimycins (TNMs) are a group of 10-membered anthraquinone-fused enediynes, newly discovered from Streptomyces sp. CB03234. Among them, TNM-A and TNM-D have exhibited excellent antitumor performances and could be exploited as very promising warheads for the development of anticancer antibody-drug conjugates (ADCs). However, their low titers, especially TNM-D, have severely limited following progress. Therefore, the streptomycin-induced ribosome engineering was adopted in this work for strain improvement of CB03234, and a TNMs high producer S. sp. CB03234-S with the K43N mutation at 30S ribosomal protein S12 was successfully screened out. Subsequent media Optimization revealed the essential effects of iodide and copper ion on the production of TNMs, while the substitution of nitrogen source could evidently promote the accumulation of TNM-D, and the ratio of produced TNM-A and TNM-D was responsive to the change of carbon and nitrogen ratio in the medium. Further amelioration of the pH control in scaled up 25 L Fermentation increased the average titers of TNM-A and TNM-D up to 13.7 ± 0.3 and 19.2 ± 0.4 mg/L, respectively. The achieved over 45-fold titer improvement of TNM-A, and 109-fold total titer improvement of TNM-A and TNM-D enabled the efficient purification of over 200 mg of each target molecule from 25 L Fermentation. Our efforts have demonstrated a practical strategy for titer improvement of anthraquinone-fused enediynes and set up a solid base for the pilot scale production and preclinical studies of TNMs to expedite the future development of anticancer ADC drugs.
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ribosome engineering and Fermentation Optimization leads to overproduction of tiancimycin a a new enediyne natural product from streptomyces sp cb03234
Journal of Industrial Microbiology & Biotechnology, 2018Co-Authors: Zilong Wang, Yanwen Duan, Dong Yang, Ben Shen, Yong HuangAbstract:Tiancimycin (TNM) A, a recently discovered enediyne natural product from Streptomyces sp. CB03234, showed rapid and complete killing of cancer cells and could be used as a payload in antibody drug conjugates. The low yield of TNM A in the wild-type strain promoted us to use ribosome engineering and Fermentation Optimization for its yield improvement. The Streptomyces sp. CB03234-R-16 mutant strain with a L422P mutation in RpoB, the RNA polymerase β-subunit, was obtained from the rifamycin-resistant screening. After Fermentation Optimization, the titers of TNM A in Streptomyces sp. CB03234-R-16 reached to 22.5 ± 3.1 mg L−1 in shaking flasks, and 13 ± 1 mg L−1 in 15 L fermentors, which were at least 40-fold higher than that in the wild-type strain (~ 0.3 mg L−1). Quantitative real-time RT-PCR revealed markedly enhanced expression of key genes encoding TNM A biosynthetic enzymes and regulators in Streptomyces sp. CB03234-R-16. Our study should greatly facilitate the future efforts to develop TNM A into a clinical anticancer drug.
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strain improvement by combined uv mutagenesis and ribosome engineering and subsequent Fermentation Optimization for enhanced 6 deoxy bleomycin z production
Applied Microbiology and Biotechnology, 2018Co-Authors: Xiangcheng Zhu, Jieqian Kong, Hu Yang, Rong Huang, Yong Huang, Dong Yang, Ben Shen, Yanwen DuanAbstract:The bleomycins (BLMs) are important clinical drugs extensively used in combination chemotherapy for the treatment of various cancers. Dose-dependent lung toxicity and the development of drug resistance have restricted their wide applications. 6′-Deoxy-BLM Z, a recently engineered BLM analogue with improved antitumor activity, has the potential to be developed into the next-generation BLM anticancer drug. However, its low titer in the recombinant strain Streptomyces flavoviridis SB9026 has hampered current efforts, which require sufficient compound, to pursue preclinical studies and subsequent clinical development. Here, we report the strain improvement by combined UV mutagenesis and ribosome engineering, as well as the Fermentation Optimization, for enhanced 6′-deoxy-BLM production. A high producer, named S. flavoviridis G-4F12, was successfully isolated, producing 6′-deoxy-BLM at above 70 mg/L under the optimized Fermentation conditions, representing a sevenfold increase in comparison with that of the original producer. These findings demonstrated the effectiveness of combined empirical breeding methods in strain improvement and set the stage for sustainable production of 6′-deoxy-BLM via pilot-scale microbial Fermentation.
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generation of high rapamycin producing strain via rational metabolic pathway based mutagenesis and further titer improvement with fed batch bioprocess Optimization
Biotechnology and Bioengineering, 2010Co-Authors: Xiangcheng Zhu, Wei Zhang, Xiyang Chen, Yanwen DuanAbstract:Rapamycin is a triene macrolide antibiotic produced by Streptomyces hygroscopicus. Besides its wide application as an effective immunosuppressive agent, other important bioactivities have made rapamycin a potential drug lead for novel pharmaceutical development. However, the low titer of rapamycin in the original producer strain limits further industrialization efforts and restricts its use for other applications. Predicated on knowledge of the metabolic pathways related to rapamycin biosynthesis in S. hygroscopicus, we have rationally designed approaches to generate a rapamycin high producer strain of S. hygroscopicus HD-04-S. These have included alleviation of glucose repression, improved tolerance towards lysine and shikimic acid, and auxotrophy of tryptophan and phenylalanine through the application of stepwise UV mutagenesis. The resultant strain produced rapamycin at 450 mg/L in the shake flask scale. These Fermentations were further scaled up in 120 and 20,000 L fermentors, respectively, at the pilot plant. Selected Fermentation factors including agitation speed, pH, and on-line supplementation were systematically evaluated. A fed-batch strategy was established to maximize rapamycin production. With these efforts, an optimized Fermentation process in the larger scale fermentor was developed. The final titer of rapamycin was 812 mg/L in the 120 L fermentor and 783 mg/L in the 20,000 L fermentor. This work highlights a high rapamycin producing strain derived by mutagenesis and subsequent screening, Fermentation Optimization of which has now made it feasible to produce rapamycin on an industrial scale by Fermentation. The strategies developed here should also be applicable to titer improvement of other important microbial natural products on an industrial scale.
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generation of high rapamycin producing strain via rational metabolic pathway based mutagenesis and further titer improvement with fed batch bioprocess Optimization
Biotechnology and Bioengineering, 2010Co-Authors: Wei Zhang, Xiyang Chen, Yanwen Duan, Hu Wu, Zhi Nan XuAbstract:Rapamycin is a triene macrolide antibiotic produced by Streptomyces hygroscopicus. Besides its wide application as an effective immunosuppressive agent, other important bioactivities have made rapamycin a potential drug lead for novel pharmaceutical development. However, the low titer of rapamycin in the original producer strain limits further industrialization efforts and restricts its use for other applications. Predicated on knowledge of the metabolic pathways related to rapamycin biosynthesis in S. hygroscopicus, we have rationally designed approaches to generate a rapamycin high producer strain of S. hygroscopicus HD-04-S. These have included alleviation of glucose repression, improved tolerance towards lysine and shikimic acid, and auxotrophy of tryptophan and phenylalanine through the application of stepwise UV mutagenesis. The resultant strain produced rapamycin at 450 mg/L in the shake flask scale. These Fermentations were further scaled up in 120 and 20,000 L fermentors, respectively, at the pilot plant. Selected Fermentation factors including agitation speed, pH, and on-line supplementation were systematically evaluated. A fed-batch strategy was established to maximize rapamycin production. With these efforts, an optimized Fermentation process in the larger scale fermentor was developed. The final titer of rapamycin was 812 mg/L in the 120 L fermentor and 783 mg/L in the 20,000 L fermentor. This work highlights a high rapamycin producing strain derived by mutagenesis and subsequent screening, Fermentation Optimization of which has now made it feasible to produce rapamycin on an industrial scale by Fermentation. The strategies developed here should also be applicable to titer improvement of other important microbial natural products on an industrial scale. Biotechnol. Bioeng. 2010;107: 506–515. © 2010 Wiley Periodicals, Inc.
Xiangcheng Zhu - One of the best experts on this subject based on the ideXlab platform.
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streptomycin induced ribosome engineering complemented with Fermentation Optimization for enhanced production of 10 membered enediynes tiancimycin a and tiancimycin d
Biotechnology and Bioengineering, 2019Co-Authors: Zhoukang Zhuang, Yanwen Duan, Rong Huang, Yong Huang, Chengzhou Jiang, Fan Zhang, Xiaohui Yan, Xiangcheng ZhuAbstract:Tiancimycins (TNMs) are a group of 10-membered anthraquinone-fused enediynes, newly discovered from Streptomyces sp. CB03234. Among them, TNM-A and TNM-D have exhibited excellent antitumor performances and could be exploited as very promising warheads for the development of anticancer antibody-drug conjugates (ADCs). However, their low titers, especially TNM-D, have severely limited following progress. Therefore, the streptomycin-induced ribosome engineering was adopted in this work for strain improvement of CB03234, and a TNMs high producer S. sp. CB03234-S with the K43N mutation at 30S ribosomal protein S12 was successfully screened out. Subsequent media Optimization revealed the essential effects of iodide and copper ion on the production of TNMs, while the substitution of nitrogen source could evidently promote the accumulation of TNM-D, and the ratio of produced TNM-A and TNM-D was responsive to the change of carbon and nitrogen ratio in the medium. Further amelioration of the pH control in scaled up 25 L Fermentation increased the average titers of TNM-A and TNM-D up to 13.7 ± 0.3 and 19.2 ± 0.4 mg/L, respectively. The achieved over 45-fold titer improvement of TNM-A, and 109-fold total titer improvement of TNM-A and TNM-D enabled the efficient purification of over 200 mg of each target molecule from 25 L Fermentation. Our efforts have demonstrated a practical strategy for titer improvement of anthraquinone-fused enediynes and set up a solid base for the pilot scale production and preclinical studies of TNMs to expedite the future development of anticancer ADC drugs.
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strain improvement by combined uv mutagenesis and ribosome engineering and subsequent Fermentation Optimization for enhanced 6 deoxy bleomycin z production
Applied Microbiology and Biotechnology, 2018Co-Authors: Xiangcheng Zhu, Jieqian Kong, Hu Yang, Rong Huang, Yong Huang, Dong Yang, Ben Shen, Yanwen DuanAbstract:The bleomycins (BLMs) are important clinical drugs extensively used in combination chemotherapy for the treatment of various cancers. Dose-dependent lung toxicity and the development of drug resistance have restricted their wide applications. 6′-Deoxy-BLM Z, a recently engineered BLM analogue with improved antitumor activity, has the potential to be developed into the next-generation BLM anticancer drug. However, its low titer in the recombinant strain Streptomyces flavoviridis SB9026 has hampered current efforts, which require sufficient compound, to pursue preclinical studies and subsequent clinical development. Here, we report the strain improvement by combined UV mutagenesis and ribosome engineering, as well as the Fermentation Optimization, for enhanced 6′-deoxy-BLM production. A high producer, named S. flavoviridis G-4F12, was successfully isolated, producing 6′-deoxy-BLM at above 70 mg/L under the optimized Fermentation conditions, representing a sevenfold increase in comparison with that of the original producer. These findings demonstrated the effectiveness of combined empirical breeding methods in strain improvement and set the stage for sustainable production of 6′-deoxy-BLM via pilot-scale microbial Fermentation.
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generation of high rapamycin producing strain via rational metabolic pathway based mutagenesis and further titer improvement with fed batch bioprocess Optimization
Biotechnology and Bioengineering, 2010Co-Authors: Xiangcheng Zhu, Wei Zhang, Xiyang Chen, Yanwen DuanAbstract:Rapamycin is a triene macrolide antibiotic produced by Streptomyces hygroscopicus. Besides its wide application as an effective immunosuppressive agent, other important bioactivities have made rapamycin a potential drug lead for novel pharmaceutical development. However, the low titer of rapamycin in the original producer strain limits further industrialization efforts and restricts its use for other applications. Predicated on knowledge of the metabolic pathways related to rapamycin biosynthesis in S. hygroscopicus, we have rationally designed approaches to generate a rapamycin high producer strain of S. hygroscopicus HD-04-S. These have included alleviation of glucose repression, improved tolerance towards lysine and shikimic acid, and auxotrophy of tryptophan and phenylalanine through the application of stepwise UV mutagenesis. The resultant strain produced rapamycin at 450 mg/L in the shake flask scale. These Fermentations were further scaled up in 120 and 20,000 L fermentors, respectively, at the pilot plant. Selected Fermentation factors including agitation speed, pH, and on-line supplementation were systematically evaluated. A fed-batch strategy was established to maximize rapamycin production. With these efforts, an optimized Fermentation process in the larger scale fermentor was developed. The final titer of rapamycin was 812 mg/L in the 120 L fermentor and 783 mg/L in the 20,000 L fermentor. This work highlights a high rapamycin producing strain derived by mutagenesis and subsequent screening, Fermentation Optimization of which has now made it feasible to produce rapamycin on an industrial scale by Fermentation. The strategies developed here should also be applicable to titer improvement of other important microbial natural products on an industrial scale.
Wei Zhang - One of the best experts on this subject based on the ideXlab platform.
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generation of high rapamycin producing strain via rational metabolic pathway based mutagenesis and further titer improvement with fed batch bioprocess Optimization
Biotechnology and Bioengineering, 2010Co-Authors: Xiangcheng Zhu, Wei Zhang, Xiyang Chen, Yanwen DuanAbstract:Rapamycin is a triene macrolide antibiotic produced by Streptomyces hygroscopicus. Besides its wide application as an effective immunosuppressive agent, other important bioactivities have made rapamycin a potential drug lead for novel pharmaceutical development. However, the low titer of rapamycin in the original producer strain limits further industrialization efforts and restricts its use for other applications. Predicated on knowledge of the metabolic pathways related to rapamycin biosynthesis in S. hygroscopicus, we have rationally designed approaches to generate a rapamycin high producer strain of S. hygroscopicus HD-04-S. These have included alleviation of glucose repression, improved tolerance towards lysine and shikimic acid, and auxotrophy of tryptophan and phenylalanine through the application of stepwise UV mutagenesis. The resultant strain produced rapamycin at 450 mg/L in the shake flask scale. These Fermentations were further scaled up in 120 and 20,000 L fermentors, respectively, at the pilot plant. Selected Fermentation factors including agitation speed, pH, and on-line supplementation were systematically evaluated. A fed-batch strategy was established to maximize rapamycin production. With these efforts, an optimized Fermentation process in the larger scale fermentor was developed. The final titer of rapamycin was 812 mg/L in the 120 L fermentor and 783 mg/L in the 20,000 L fermentor. This work highlights a high rapamycin producing strain derived by mutagenesis and subsequent screening, Fermentation Optimization of which has now made it feasible to produce rapamycin on an industrial scale by Fermentation. The strategies developed here should also be applicable to titer improvement of other important microbial natural products on an industrial scale.
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generation of high rapamycin producing strain via rational metabolic pathway based mutagenesis and further titer improvement with fed batch bioprocess Optimization
Biotechnology and Bioengineering, 2010Co-Authors: Wei Zhang, Xiyang Chen, Yanwen Duan, Hu Wu, Zhi Nan XuAbstract:Rapamycin is a triene macrolide antibiotic produced by Streptomyces hygroscopicus. Besides its wide application as an effective immunosuppressive agent, other important bioactivities have made rapamycin a potential drug lead for novel pharmaceutical development. However, the low titer of rapamycin in the original producer strain limits further industrialization efforts and restricts its use for other applications. Predicated on knowledge of the metabolic pathways related to rapamycin biosynthesis in S. hygroscopicus, we have rationally designed approaches to generate a rapamycin high producer strain of S. hygroscopicus HD-04-S. These have included alleviation of glucose repression, improved tolerance towards lysine and shikimic acid, and auxotrophy of tryptophan and phenylalanine through the application of stepwise UV mutagenesis. The resultant strain produced rapamycin at 450 mg/L in the shake flask scale. These Fermentations were further scaled up in 120 and 20,000 L fermentors, respectively, at the pilot plant. Selected Fermentation factors including agitation speed, pH, and on-line supplementation were systematically evaluated. A fed-batch strategy was established to maximize rapamycin production. With these efforts, an optimized Fermentation process in the larger scale fermentor was developed. The final titer of rapamycin was 812 mg/L in the 120 L fermentor and 783 mg/L in the 20,000 L fermentor. This work highlights a high rapamycin producing strain derived by mutagenesis and subsequent screening, Fermentation Optimization of which has now made it feasible to produce rapamycin on an industrial scale by Fermentation. The strategies developed here should also be applicable to titer improvement of other important microbial natural products on an industrial scale. Biotechnol. Bioeng. 2010;107: 506–515. © 2010 Wiley Periodicals, Inc.
Jinyan Zhou - One of the best experts on this subject based on the ideXlab platform.
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the artificial neural network approach based on uniform design to optimize the fed batch Fermentation condition application to the production of iturin a
Microbial Cell Factories, 2014Co-Authors: Wenjing Peng, Juan Zhong, Jie Yang, Tan Xu, Song Xiao, Jinyan ZhouAbstract:Iturin A is a potential lipopeptide antibiotic produced by Bacillus subtilis. Optimization of iturin A yield by adding various concentrations of asparagine (Asn), glutamic acid (Glu) and proline (Pro) during the fed-batch Fermentation process was studied using an artificial neural network-genetic algorithm (ANN-GA) and uniform design (UD). Here, ANN-GA based on the UD data was used for the first time to analyze the fed-batch Fermentation process. The ANN-GA and UD methodologies were compared based on their fitting ability, prediction and generalization capacity and sensitivity analysis. The ANN model based on the UD data performed well on minimal statistical designed experimental number and the optimum iturin A yield was 13364.5 ± 271.3 U/mL compared with a yield of 9929.0 ± 280.9 U/mL for the control (batch Fermentation without adding the amino acids). The root-mean-square-error for the ANN model with the training set and test set was 4.84 and 273.58 respectively, which was more than two times better than that for the UD model (32.21 and 483.12). The correlation coefficient for the ANN model with training and test sets was 100% and 92.62%, respectively (compared with 99.86% and 78.58% for UD). The error% for ANN with the training and test sets was 0.093 and 2.19 respectively (compared with 0.26 and 4.15 for UD). The sensitivity analysis of both methods showed the comparable results. The predictive error of the optimal iturin A yield for ANN-GA and UD was 0.8% and 2.17%, respectively. The satisfactory fitting and predicting accuracy of ANN indicated that ANN worked well with the UD data. Through ANN-GA, the iturin A yield was significantly increased by 34.6%. The fitness, prediction, and generalization capacities of the ANN model were better than those of the UD model. Further, although UD could get the insight information between variables directly, ANN was also demonstrated to be efficient in the sensitivity analysis. The results of these comparisons indicated that ANN could be a better alternative way for Fermentation Optimization with limited number of experiments.
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the artificial neural network approach based on uniform design to optimize the fed batch Fermentation condition application to the production of iturin a
Microbial Cell Factories, 2014Co-Authors: Wenjing Peng, Juan Zhong, Jie Yang, Song Xiao, Jinyan Zhou, Yanli Ren, Hong TanAbstract:Background: Iturin A is a potential lipopeptide antibiotic produced by Bacillus subtilis. Optimization of iturin A yield by adding various concentrations of asparagine (Asn), glutamic acid (Glu) and proline (Pro) during the fed-batch Fermentation process was studied using an artificial neural network-genetic algorithm (ANN-GA) and uniform design (UD). Here, ANN-GA based on the UD data was used for the first time to analyze the fed-batch Fermentation process. The ANN-GA and UD methodologies were compared based on their fitting ability, prediction and generalization capacity and sensitivity analysis. Results: The ANN model based on the UD data performed well on minimal statistical designed experimental number and the optimum iturin A yield was 13364.5 ± 271.3 U/mL compared with a yield of 9929.0 ± 280.9 U/mL for the control (batch Fermentation without adding the amino acids). The root-mean-square-error for the ANN model with the training set and test set was 4.84 and 273.58 respectively, which was more than two times better than that for the UD model (32.21 and 483.12). The correlation coefficient for the ANN model with training and test sets was 100% and 92.62%, respectively (compared with 99.86% and 78.58% for UD). The error% for ANN with the training and test sets was 0.093 and 2.19 respectively (compared with 0.26 and 4.15 for UD). The sensitivity analysis of both methods showed the comparable results. The predictive error of the optimal iturin A yield for ANN-GA and UD was 0.8% and 2.17%, respectively. Conclusions: The satisfactory fitting and predicting accuracy of ANN indicated that ANN worked well with the UD data. Through ANN-GA, the iturin A yield was significantly increased by 34.6%. The fitness, prediction, and generalization capacities of the ANN model were better than those of the UD model. Further, although UD could get the insight information between variables directly, ANN was also demonstrated to be efficient in the sensitivity analysis. The results of these comparisons indicated that ANN could be a better alternative way for Fermentation Optimization with limited number of experiments.
Zhi Nan Xu - One of the best experts on this subject based on the ideXlab platform.
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generation of high rapamycin producing strain via rational metabolic pathway based mutagenesis and further titer improvement with fed batch bioprocess Optimization
Biotechnology and Bioengineering, 2010Co-Authors: Wei Zhang, Xiyang Chen, Yanwen Duan, Hu Wu, Zhi Nan XuAbstract:Rapamycin is a triene macrolide antibiotic produced by Streptomyces hygroscopicus. Besides its wide application as an effective immunosuppressive agent, other important bioactivities have made rapamycin a potential drug lead for novel pharmaceutical development. However, the low titer of rapamycin in the original producer strain limits further industrialization efforts and restricts its use for other applications. Predicated on knowledge of the metabolic pathways related to rapamycin biosynthesis in S. hygroscopicus, we have rationally designed approaches to generate a rapamycin high producer strain of S. hygroscopicus HD-04-S. These have included alleviation of glucose repression, improved tolerance towards lysine and shikimic acid, and auxotrophy of tryptophan and phenylalanine through the application of stepwise UV mutagenesis. The resultant strain produced rapamycin at 450 mg/L in the shake flask scale. These Fermentations were further scaled up in 120 and 20,000 L fermentors, respectively, at the pilot plant. Selected Fermentation factors including agitation speed, pH, and on-line supplementation were systematically evaluated. A fed-batch strategy was established to maximize rapamycin production. With these efforts, an optimized Fermentation process in the larger scale fermentor was developed. The final titer of rapamycin was 812 mg/L in the 120 L fermentor and 783 mg/L in the 20,000 L fermentor. This work highlights a high rapamycin producing strain derived by mutagenesis and subsequent screening, Fermentation Optimization of which has now made it feasible to produce rapamycin on an industrial scale by Fermentation. The strategies developed here should also be applicable to titer improvement of other important microbial natural products on an industrial scale. Biotechnol. Bioeng. 2010;107: 506–515. © 2010 Wiley Periodicals, Inc.