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      Modular engineering to increase intracellular NAD(H/+) promotes rate of extracellular electron transfer of Shewanella oneidensis

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          Abstract

          The slow rate of extracellular electron transfer (EET) of electroactive microorganisms remains a primary bottleneck that restricts the practical applications of bioelectrochemical systems. Intracellular NAD(H/ +) (i.e., the total level of NADH and NAD +) is a crucial source of the intracellular electron pool from which intracellular electrons are transferred to extracellular electron acceptors via EET pathways. However, how the total level of intracellular NAD(H/ +) impacts the EET rate in Shewanella oneidensis has not been established. Here, we use a modular synthetic biology strategy to redirect metabolic flux towards NAD + biosynthesis via three modules: de novo, salvage, and universal biosynthesis modules in S. oneidensis MR-1. The results demonstrate that an increase in intracellular NAD(H/ +) results in the transfer of more electrons from the increased oxidation of the electron donor to the EET pathways of S. oneidensis, thereby enhancing intracellular electron flux and the EET rate.

          Abstract

          A bottleneck for the application of bioelectrochemical systems is the slow rate of extracellular electron transfer. Here the authors use a synthetic biology approach to redirect metabolic flux to NAD + biosynthesis, which enhances the intracellular electron flux and the extracellular electron transfer rate.

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          Most cited references 66

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          Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method.

           K Livak,  T Schmittgen (2001)
          The two most commonly used methods to analyze data from real-time, quantitative PCR experiments are absolute quantification and relative quantification. Absolute quantification determines the input copy number, usually by relating the PCR signal to a standard curve. Relative quantification relates the PCR signal of the target transcript in a treatment group to that of another sample such as an untreated control. The 2(-Delta Delta C(T)) method is a convenient way to analyze the relative changes in gene expression from real-time quantitative PCR experiments. The purpose of this report is to present the derivation, assumptions, and applications of the 2(-Delta Delta C(T)) method. In addition, we present the derivation and applications of two variations of the 2(-Delta Delta C(T)) method that may be useful in the analysis of real-time, quantitative PCR data. Copyright 2001 Elsevier Science (USA).
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            Tricine-SDS-PAGE.

            Tricine-SDS-PAGE is commonly used to separate proteins in the mass range 1-100 kDa. It is the preferred electrophoretic system for the resolution of proteins smaller than 30 kDa. The concentrations of acrylamide used in the gels are lower than in other electrophoretic systems. These lower concentrations facilitate electroblotting, which is particularly crucial for hydrophobic proteins. Tricine-SDS-PAGE is also used preferentially for doubled SDS-PAGE (dSDS-PAGE), a proteomic tool used to isolate extremely hydrophobic proteins for mass spectrometric identification, and it offers advantages for resolution of the second dimension after blue-native PAGE (BN-PAGE) and clear-native PAGE (CN-PAGE). Here I describe a protocol for Tricine-SDS-PAGE, which includes efficient methods for Coomassie blue or silver staining and electroblotting, thereby increasing the versatility of the approach. This protocol can be completed in 1-2 d.
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              Exoelectrogenic bacteria that power microbial fuel cells.

               Bruce Logan (2009)
              There has been an increase in recent years in the number of reports of microorganisms that can generate electrical current in microbial fuel cells. Although many new strains have been identified, few strains individually produce power densities as high as strains from mixed communities. Enriched anodic biofilms have generated power densities as high as 6.9 W per m(2) (projected anode area), and therefore are approaching theoretical limits. To understand bacterial versatility in mechanisms used for current generation, this Progress article explores the underlying reasons for exocellular electron transfer, including cellular respiration and possible cell-cell communication.
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                Author and article information

                Affiliations
                [1 ]ISNI 0000 0004 1761 2484, GRID grid.33763.32, Key Laboratory of Systems Bioengineering (Ministry of Education), SynBio Research Platform, Collaborative Innovation Centre of Chemical Science and Engineering, School of Chemical Engineering and Technology, , Tianjin University, ; Tianjin, 300072 PR China
                [2 ]ISNI 0000 0001 0373 6302, GRID grid.428986.9, State Key Laboratory of Marine Resource Utilization in South China Sea, College of Information Science & Technology, , Hainan University, ; Haikou, 570228 PR China
                [3 ]ISNI 0000 0004 0368 8293, GRID grid.16821.3c, State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, , Shanghai Jiao Tong University, ; Shanghai, 200240 PR China
                [4 ]Department of Biological Sciences and Technology, School of Environmental Studies, China University of Geoscience in Wuhan, Wuhan, 430074 Hubei PR China
                Contributors
                hsong@tju.edu.cn
                Journal
                Nat Commun
                Nat Commun
                Nature Communications
                Nature Publishing Group UK (London )
                2041-1723
                7 September 2018
                7 September 2018
                2018
                : 9
                5995
                10.1038/s41467-018-05995-8
                6128845
                30194293
                © The Author(s) 2018

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

                Funding
                Funded by: the Open Project Program of the State Key Lab of Marine Resource Utilization in South China Sea (Hainan University) granted (No. 2016010)
                Funded by: the State Key Laboratory of Microbial Metabolism (Shanghai Jiao Tong University, MMLKF 17-12)
                Funded by: the National Natural Science Foundation of China (NSFC 41630318, 41772363)
                Funded by: the National Natural Science Foundation of China (NSFC 21376174, 21621004), the National Basic Research Program of China (“973” Program: 2014CB745103), the Open Project Program of the State Key Lab of Marine Resource Utilization in South China Sea (Hainan University) granted (No. 2016010), and the State Key Laboratory of Microbial Metabolism (Shanghai Jiao Tong University, MMLKF 17-12).
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