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Combinatorial optimization of CO sub(2) transport and fixation to improve succinate production by promoter engineering
To balance the flux of an engineered metabolic pathway to achieve high yield of target product is a major challenge in metabolic engineering. In previous work, the collaborative regulation of CO sub(2) transport and fixation was investigated with co-overexpressing exogenous genes regulating both CO...
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Published in: | Biotechnology and bioengineering 2016-07, Vol.113 (7), p.1531-1541 |
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creator | Yu, Jun-Han Zhu, Li-Wen Xia, Shi-Tao Li, Hong-Mei Tang, Ya-Ling Liang, Xin-Hua Chen, Tao Tang, Ya-Jie |
description | To balance the flux of an engineered metabolic pathway to achieve high yield of target product is a major challenge in metabolic engineering. In previous work, the collaborative regulation of CO sub(2) transport and fixation was investigated with co-overexpressing exogenous genes regulating both CO sub(2) transport (sbtA and bicA) and PEP carboxylation (phosphoenolpyruvate (PEP) carboxylase (ppc) and carboxykinase (pck)) under trc promoter in Escherichia coli for succinate biosynthesis. For balancing metabolic flux to maximize succinate titer, a combinatorial optimization strategy to fine-tuning CO sub(2) transport and fixation process was implemented by promoter engineering in this study. Firstly, based on the energy matrix a synthetic promoter library containing 20 rationally designed promoters with strengths ranging from 0.8% to 100% compared with the widely used trc promoter was generated. Evaluations of rfp and cat reporter genes provided evidence that the synthetic promoters were stably and had certain applicability. Secondly, four designed promoters with different strengths were used for combinatorial assembly of single CO sub(2) transport gene (sbtA or bicA) and single CO sub(2) fixation gene (ppc or pck) expression. Three combinations, such as Tang1519 (P sub(4)-bicA+pP sub(19)-pck), Tang1522 (P sub(4)-sbtA+P sub(4)-ppc), Tang1523 (P sub(4)-sbtA+P sub(17)-ppc) with a more than 10% increase in succinate production were screened in bioreactor. Finally, based on the above results, co-expression of the four transport and fixation genes were further investigated. Co-expression of sbtA, bicA, and ppc with weak promoter P sub(4) and pck with strong promoter P sub(19) (AFP111/pT-P sub(4)-bicA-P sub(4)-sbtA+pACYC-P sub(19)-pck-P sub(4)-ppc) provided the best succinate production among all the combinations. The highest succinate production of 89.4g/L was 37.5% higher than that obtained with empty vector control. This work significantly enhanced succinate production through combinatorial optimization of CO sub(2) transport and fixation. The promoter engineering and combinatorial optimization strategies used herein represents a powerful approach to tailor-making metabolic pathways for the production of other industrially important chemicals. Biotechnol. Bioeng. 2016; 113: 1531-1541. For balancing metabolic flux to maximize succinate titer, a combinatorial optimization strategy to fine-tuning CO sub(2) transport and fixation process was implemented by promoter |
doi_str_mv | 10.1002/bit.25927 |
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In previous work, the collaborative regulation of CO sub(2) transport and fixation was investigated with co-overexpressing exogenous genes regulating both CO sub(2) transport (sbtA and bicA) and PEP carboxylation (phosphoenolpyruvate (PEP) carboxylase (ppc) and carboxykinase (pck)) under trc promoter in Escherichia coli for succinate biosynthesis. For balancing metabolic flux to maximize succinate titer, a combinatorial optimization strategy to fine-tuning CO sub(2) transport and fixation process was implemented by promoter engineering in this study. Firstly, based on the energy matrix a synthetic promoter library containing 20 rationally designed promoters with strengths ranging from 0.8% to 100% compared with the widely used trc promoter was generated. Evaluations of rfp and cat reporter genes provided evidence that the synthetic promoters were stably and had certain applicability. Secondly, four designed promoters with different strengths were used for combinatorial assembly of single CO sub(2) transport gene (sbtA or bicA) and single CO sub(2) fixation gene (ppc or pck) expression. Three combinations, such as Tang1519 (P sub(4)-bicA+pP sub(19)-pck), Tang1522 (P sub(4)-sbtA+P sub(4)-ppc), Tang1523 (P sub(4)-sbtA+P sub(17)-ppc) with a more than 10% increase in succinate production were screened in bioreactor. Finally, based on the above results, co-expression of the four transport and fixation genes were further investigated. Co-expression of sbtA, bicA, and ppc with weak promoter P sub(4) and pck with strong promoter P sub(19) (AFP111/pT-P sub(4)-bicA-P sub(4)-sbtA+pACYC-P sub(19)-pck-P sub(4)-ppc) provided the best succinate production among all the combinations. The highest succinate production of 89.4g/L was 37.5% higher than that obtained with empty vector control. This work significantly enhanced succinate production through combinatorial optimization of CO sub(2) transport and fixation. The promoter engineering and combinatorial optimization strategies used herein represents a powerful approach to tailor-making metabolic pathways for the production of other industrially important chemicals. Biotechnol. Bioeng. 2016; 113: 1531-1541. For balancing metabolic flux to maximize succinate titer, a combinatorial optimization strategy to fine-tuning CO sub(2) transport and fixation process was implemented by promoter engineering in this study. Four designed promoters with different strengths were used for combinatorial assembly of CO sub(2) transport genes (sbtA and bicA) and CO sub(2) fixation genes (ppc and pck) expression. Co-expression of sbtA, bicA, and ppc with weak promoter P sub(4) and pck with strong promoter P sub(19) provided the best succinate production among all the combination.</description><identifier>ISSN: 0006-3592</identifier><identifier>EISSN: 1097-0290</identifier><identifier>DOI: 10.1002/bit.25927</identifier><language>eng</language><subject>Carbon capture and storage ; Carbon dioxide ; Combinatorial analysis ; Escherichia coli ; Fixation ; Gene expression ; Genes ; Optimization ; Transport</subject><ispartof>Biotechnology and bioengineering, 2016-07, Vol.113 (7), p.1531-1541</ispartof><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids></links><search><creatorcontrib>Yu, Jun-Han</creatorcontrib><creatorcontrib>Zhu, Li-Wen</creatorcontrib><creatorcontrib>Xia, Shi-Tao</creatorcontrib><creatorcontrib>Li, Hong-Mei</creatorcontrib><creatorcontrib>Tang, Ya-Ling</creatorcontrib><creatorcontrib>Liang, Xin-Hua</creatorcontrib><creatorcontrib>Chen, Tao</creatorcontrib><creatorcontrib>Tang, Ya-Jie</creatorcontrib><title>Combinatorial optimization of CO sub(2) transport and fixation to improve succinate production by promoter engineering</title><title>Biotechnology and bioengineering</title><description>To balance the flux of an engineered metabolic pathway to achieve high yield of target product is a major challenge in metabolic engineering. In previous work, the collaborative regulation of CO sub(2) transport and fixation was investigated with co-overexpressing exogenous genes regulating both CO sub(2) transport (sbtA and bicA) and PEP carboxylation (phosphoenolpyruvate (PEP) carboxylase (ppc) and carboxykinase (pck)) under trc promoter in Escherichia coli for succinate biosynthesis. For balancing metabolic flux to maximize succinate titer, a combinatorial optimization strategy to fine-tuning CO sub(2) transport and fixation process was implemented by promoter engineering in this study. Firstly, based on the energy matrix a synthetic promoter library containing 20 rationally designed promoters with strengths ranging from 0.8% to 100% compared with the widely used trc promoter was generated. Evaluations of rfp and cat reporter genes provided evidence that the synthetic promoters were stably and had certain applicability. Secondly, four designed promoters with different strengths were used for combinatorial assembly of single CO sub(2) transport gene (sbtA or bicA) and single CO sub(2) fixation gene (ppc or pck) expression. Three combinations, such as Tang1519 (P sub(4)-bicA+pP sub(19)-pck), Tang1522 (P sub(4)-sbtA+P sub(4)-ppc), Tang1523 (P sub(4)-sbtA+P sub(17)-ppc) with a more than 10% increase in succinate production were screened in bioreactor. Finally, based on the above results, co-expression of the four transport and fixation genes were further investigated. Co-expression of sbtA, bicA, and ppc with weak promoter P sub(4) and pck with strong promoter P sub(19) (AFP111/pT-P sub(4)-bicA-P sub(4)-sbtA+pACYC-P sub(19)-pck-P sub(4)-ppc) provided the best succinate production among all the combinations. The highest succinate production of 89.4g/L was 37.5% higher than that obtained with empty vector control. This work significantly enhanced succinate production through combinatorial optimization of CO sub(2) transport and fixation. The promoter engineering and combinatorial optimization strategies used herein represents a powerful approach to tailor-making metabolic pathways for the production of other industrially important chemicals. Biotechnol. Bioeng. 2016; 113: 1531-1541. For balancing metabolic flux to maximize succinate titer, a combinatorial optimization strategy to fine-tuning CO sub(2) transport and fixation process was implemented by promoter engineering in this study. Four designed promoters with different strengths were used for combinatorial assembly of CO sub(2) transport genes (sbtA and bicA) and CO sub(2) fixation genes (ppc and pck) expression. Co-expression of sbtA, bicA, and ppc with weak promoter P sub(4) and pck with strong promoter P sub(19) provided the best succinate production among all the combination.</description><subject>Carbon capture and storage</subject><subject>Carbon dioxide</subject><subject>Combinatorial analysis</subject><subject>Escherichia coli</subject><subject>Fixation</subject><subject>Gene expression</subject><subject>Genes</subject><subject>Optimization</subject><subject>Transport</subject><issn>0006-3592</issn><issn>1097-0290</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2016</creationdate><recordtype>article</recordtype><recordid>eNqNjslOwzAURS0EEqWw4A-8LIuAh3jIEkVMUqVuuq8c57kySuwQOxXw9aSUD2D17tE9unoI3VJyTwlhD43P90xUTJ2hBSWVKgiryDlaEEJkwefiEl2l9D6j0lIu0KGOfeODyXH0psNxyL733yb7GHB0uN7gNDUrdofzaEIa4pixCS12_vPk5Ih9P4zxALNo7XEJ8MztZH_75utIfcwwYgh7HwBGH_bX6MKZLsHN312i7fPTtn4t1puXt_pxXQxSqkKUyjFuQShLpBDUla1WRjHFhbNtAxS04dY46jS0x9Cy0lQaVCVKQYTkS7Q6zc4_fEyQ8q73yULXmQBxSjuq2WxWtFL_UImWXJaM8R8zsm7n</recordid><startdate>20160701</startdate><enddate>20160701</enddate><creator>Yu, Jun-Han</creator><creator>Zhu, Li-Wen</creator><creator>Xia, Shi-Tao</creator><creator>Li, Hong-Mei</creator><creator>Tang, Ya-Ling</creator><creator>Liang, Xin-Hua</creator><creator>Chen, Tao</creator><creator>Tang, Ya-Jie</creator><scope>7QO</scope><scope>8FD</scope><scope>FR3</scope><scope>P64</scope><scope>7U5</scope><scope>F28</scope><scope>L7M</scope></search><sort><creationdate>20160701</creationdate><title>Combinatorial optimization of CO sub(2) transport and fixation to improve succinate production by promoter engineering</title><author>Yu, Jun-Han ; Zhu, Li-Wen ; Xia, Shi-Tao ; Li, Hong-Mei ; Tang, Ya-Ling ; Liang, Xin-Hua ; Chen, Tao ; Tang, Ya-Jie</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-p667-547f23ce57c06551f4d87a72735fcdbe1e8a3caf1f8ed3cafd24a98e795450563</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2016</creationdate><topic>Carbon capture and storage</topic><topic>Carbon dioxide</topic><topic>Combinatorial analysis</topic><topic>Escherichia coli</topic><topic>Fixation</topic><topic>Gene expression</topic><topic>Genes</topic><topic>Optimization</topic><topic>Transport</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Yu, Jun-Han</creatorcontrib><creatorcontrib>Zhu, Li-Wen</creatorcontrib><creatorcontrib>Xia, Shi-Tao</creatorcontrib><creatorcontrib>Li, Hong-Mei</creatorcontrib><creatorcontrib>Tang, Ya-Ling</creatorcontrib><creatorcontrib>Liang, Xin-Hua</creatorcontrib><creatorcontrib>Chen, Tao</creatorcontrib><creatorcontrib>Tang, Ya-Jie</creatorcontrib><collection>Biotechnology Research Abstracts</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Solid State and Superconductivity Abstracts</collection><collection>ANTE: Abstracts in New Technology & Engineering</collection><collection>Advanced Technologies Database with Aerospace</collection><jtitle>Biotechnology and bioengineering</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Yu, Jun-Han</au><au>Zhu, Li-Wen</au><au>Xia, Shi-Tao</au><au>Li, Hong-Mei</au><au>Tang, Ya-Ling</au><au>Liang, Xin-Hua</au><au>Chen, Tao</au><au>Tang, Ya-Jie</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Combinatorial optimization of CO sub(2) transport and fixation to improve succinate production by promoter engineering</atitle><jtitle>Biotechnology and bioengineering</jtitle><date>2016-07-01</date><risdate>2016</risdate><volume>113</volume><issue>7</issue><spage>1531</spage><epage>1541</epage><pages>1531-1541</pages><issn>0006-3592</issn><eissn>1097-0290</eissn><abstract>To balance the flux of an engineered metabolic pathway to achieve high yield of target product is a major challenge in metabolic engineering. In previous work, the collaborative regulation of CO sub(2) transport and fixation was investigated with co-overexpressing exogenous genes regulating both CO sub(2) transport (sbtA and bicA) and PEP carboxylation (phosphoenolpyruvate (PEP) carboxylase (ppc) and carboxykinase (pck)) under trc promoter in Escherichia coli for succinate biosynthesis. For balancing metabolic flux to maximize succinate titer, a combinatorial optimization strategy to fine-tuning CO sub(2) transport and fixation process was implemented by promoter engineering in this study. Firstly, based on the energy matrix a synthetic promoter library containing 20 rationally designed promoters with strengths ranging from 0.8% to 100% compared with the widely used trc promoter was generated. Evaluations of rfp and cat reporter genes provided evidence that the synthetic promoters were stably and had certain applicability. Secondly, four designed promoters with different strengths were used for combinatorial assembly of single CO sub(2) transport gene (sbtA or bicA) and single CO sub(2) fixation gene (ppc or pck) expression. Three combinations, such as Tang1519 (P sub(4)-bicA+pP sub(19)-pck), Tang1522 (P sub(4)-sbtA+P sub(4)-ppc), Tang1523 (P sub(4)-sbtA+P sub(17)-ppc) with a more than 10% increase in succinate production were screened in bioreactor. Finally, based on the above results, co-expression of the four transport and fixation genes were further investigated. Co-expression of sbtA, bicA, and ppc with weak promoter P sub(4) and pck with strong promoter P sub(19) (AFP111/pT-P sub(4)-bicA-P sub(4)-sbtA+pACYC-P sub(19)-pck-P sub(4)-ppc) provided the best succinate production among all the combinations. The highest succinate production of 89.4g/L was 37.5% higher than that obtained with empty vector control. This work significantly enhanced succinate production through combinatorial optimization of CO sub(2) transport and fixation. The promoter engineering and combinatorial optimization strategies used herein represents a powerful approach to tailor-making metabolic pathways for the production of other industrially important chemicals. Biotechnol. Bioeng. 2016; 113: 1531-1541. For balancing metabolic flux to maximize succinate titer, a combinatorial optimization strategy to fine-tuning CO sub(2) transport and fixation process was implemented by promoter engineering in this study. Four designed promoters with different strengths were used for combinatorial assembly of CO sub(2) transport genes (sbtA and bicA) and CO sub(2) fixation genes (ppc and pck) expression. Co-expression of sbtA, bicA, and ppc with weak promoter P sub(4) and pck with strong promoter P sub(19) provided the best succinate production among all the combination.</abstract><doi>10.1002/bit.25927</doi><tpages>11</tpages></addata></record> |
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subjects | Carbon capture and storage Carbon dioxide Combinatorial analysis Escherichia coli Fixation Gene expression Genes Optimization Transport |
title | Combinatorial optimization of CO sub(2) transport and fixation to improve succinate production by promoter engineering |
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