Table_1_Proteomics Reveal the Effect of Exogenous Electrons on Electroactive Escherichia coli.DOCX

Microbial cells utilizing electricity to produce high-value fuels and chemicals are the foundation of the biocathodic bioelectrochemical system. However, molecular mechanisms of electron transfer and utilization have not been elucidated. In this work, Escherichia coli engineered by introducing the Mtr pathway from Shewanella oneidensis exhibited stronger electrochemical activity than control and could utilize exogenous electrons to stimulate metabolite profiles and boost succinate production in the bioelectrochemical system. Proteomic analysis and real-time PCR were performed to investigate the effect of exogenous electrons on electroactive E. coli. Bioinformatics analysis suggested that the proteins of molecular function associated with oxidoreductase activity, 4 iron, 4 sulfur([4Fe-4S]) cluster binding, iron-sulfur cluster binding, and metal cluster binding were positively affected by exogenous electrons. Moreover, mapping to the Kyoto Encyclopedia of Genes and Genomes pathway database showed that the up-regulated proteins were mainly involved in metabolic pathways of tricarboxylic acid cycle, pyruvate metabolism, and nitrogen metabolism pathway, providing support for the metabolic balance of microbial cells shifting toward reduced end-products due to electron utilization. Using a biochemical method, the ompF-overexpressed strain was employed to investigate the function of the channel protein. These findings provided a theoretical basis for further improving electron transfer and utilization efficiency, and contributed to the potential applications of the bioelectrochemical system.</p

Table_2_Proteomics Reveal the Effect of Exogenous Electrons on Electroactive Escherichia coli.DOCX

Microbial cells utilizing electricity to produce high-value fuels and chemicals are the foundation of the biocathodic bioelectrochemical system. However, molecular mechanisms of electron transfer and utilization have not been elucidated. In this work, Escherichia coli engineered by introducing the Mtr pathway from Shewanella oneidensis exhibited stronger electrochemical activity than control and could utilize exogenous electrons to stimulate metabolite profiles and boost succinate production in the bioelectrochemical system. Proteomic analysis and real-time PCR were performed to investigate the effect of exogenous electrons on electroactive E. coli. Bioinformatics analysis suggested that the proteins of molecular function associated with oxidoreductase activity, 4 iron, 4 sulfur([4Fe-4S]) cluster binding, iron-sulfur cluster binding, and metal cluster binding were positively affected by exogenous electrons. Moreover, mapping to the Kyoto Encyclopedia of Genes and Genomes pathway database showed that the up-regulated proteins were mainly involved in metabolic pathways of tricarboxylic acid cycle, pyruvate metabolism, and nitrogen metabolism pathway, providing support for the metabolic balance of microbial cells shifting toward reduced end-products due to electron utilization. Using a biochemical method, the ompF-overexpressed strain was employed to investigate the function of the channel protein. These findings provided a theoretical basis for further improving electron transfer and utilization efficiency, and contributed to the potential applications of the bioelectrochemical system.</p

Table_4_Proteomics Reveal the Effect of Exogenous Electrons on Electroactive Escherichia coli.docx

Microbial cells utilizing electricity to produce high-value fuels and chemicals are the foundation of the biocathodic bioelectrochemical system. However, molecular mechanisms of electron transfer and utilization have not been elucidated. In this work, Escherichia coli engineered by introducing the Mtr pathway from Shewanella oneidensis exhibited stronger electrochemical activity than control and could utilize exogenous electrons to stimulate metabolite profiles and boost succinate production in the bioelectrochemical system. Proteomic analysis and real-time PCR were performed to investigate the effect of exogenous electrons on electroactive E. coli. Bioinformatics analysis suggested that the proteins of molecular function associated with oxidoreductase activity, 4 iron, 4 sulfur([4Fe-4S]) cluster binding, iron-sulfur cluster binding, and metal cluster binding were positively affected by exogenous electrons. Moreover, mapping to the Kyoto Encyclopedia of Genes and Genomes pathway database showed that the up-regulated proteins were mainly involved in metabolic pathways of tricarboxylic acid cycle, pyruvate metabolism, and nitrogen metabolism pathway, providing support for the metabolic balance of microbial cells shifting toward reduced end-products due to electron utilization. Using a biochemical method, the ompF-overexpressed strain was employed to investigate the function of the channel protein. These findings provided a theoretical basis for further improving electron transfer and utilization efficiency, and contributed to the potential applications of the bioelectrochemical system.</p

Table_5_Proteomics Reveal the Effect of Exogenous Electrons on Electroactive Escherichia coli.DOCX

Microbial cells utilizing electricity to produce high-value fuels and chemicals are the foundation of the biocathodic bioelectrochemical system. However, molecular mechanisms of electron transfer and utilization have not been elucidated. In this work, Escherichia coli engineered by introducing the Mtr pathway from Shewanella oneidensis exhibited stronger electrochemical activity than control and could utilize exogenous electrons to stimulate metabolite profiles and boost succinate production in the bioelectrochemical system. Proteomic analysis and real-time PCR were performed to investigate the effect of exogenous electrons on electroactive E. coli. Bioinformatics analysis suggested that the proteins of molecular function associated with oxidoreductase activity, 4 iron, 4 sulfur([4Fe-4S]) cluster binding, iron-sulfur cluster binding, and metal cluster binding were positively affected by exogenous electrons. Moreover, mapping to the Kyoto Encyclopedia of Genes and Genomes pathway database showed that the up-regulated proteins were mainly involved in metabolic pathways of tricarboxylic acid cycle, pyruvate metabolism, and nitrogen metabolism pathway, providing support for the metabolic balance of microbial cells shifting toward reduced end-products due to electron utilization. Using a biochemical method, the ompF-overexpressed strain was employed to investigate the function of the channel protein. These findings provided a theoretical basis for further improving electron transfer and utilization efficiency, and contributed to the potential applications of the bioelectrochemical system.</p

Table_3_Proteomics Reveal the Effect of Exogenous Electrons on Electroactive Escherichia coli.DOCX

Microbial cells utilizing electricity to produce high-value fuels and chemicals are the foundation of the biocathodic bioelectrochemical system. However, molecular mechanisms of electron transfer and utilization have not been elucidated. In this work, Escherichia coli engineered by introducing the Mtr pathway from Shewanella oneidensis exhibited stronger electrochemical activity than control and could utilize exogenous electrons to stimulate metabolite profiles and boost succinate production in the bioelectrochemical system. Proteomic analysis and real-time PCR were performed to investigate the effect of exogenous electrons on electroactive E. coli. Bioinformatics analysis suggested that the proteins of molecular function associated with oxidoreductase activity, 4 iron, 4 sulfur([4Fe-4S]) cluster binding, iron-sulfur cluster binding, and metal cluster binding were positively affected by exogenous electrons. Moreover, mapping to the Kyoto Encyclopedia of Genes and Genomes pathway database showed that the up-regulated proteins were mainly involved in metabolic pathways of tricarboxylic acid cycle, pyruvate metabolism, and nitrogen metabolism pathway, providing support for the metabolic balance of microbial cells shifting toward reduced end-products due to electron utilization. Using a biochemical method, the ompF-overexpressed strain was employed to investigate the function of the channel protein. These findings provided a theoretical basis for further improving electron transfer and utilization efficiency, and contributed to the potential applications of the bioelectrochemical system.</p

Image_2_Proteomics Reveal the Effect of Exogenous Electrons on Electroactive Escherichia coli.pdf

Microbial cells utilizing electricity to produce high-value fuels and chemicals are the foundation of the biocathodic bioelectrochemical system. However, molecular mechanisms of electron transfer and utilization have not been elucidated. In this work, Escherichia coli engineered by introducing the Mtr pathway from Shewanella oneidensis exhibited stronger electrochemical activity than control and could utilize exogenous electrons to stimulate metabolite profiles and boost succinate production in the bioelectrochemical system. Proteomic analysis and real-time PCR were performed to investigate the effect of exogenous electrons on electroactive E. coli. Bioinformatics analysis suggested that the proteins of molecular function associated with oxidoreductase activity, 4 iron, 4 sulfur([4Fe-4S]) cluster binding, iron-sulfur cluster binding, and metal cluster binding were positively affected by exogenous electrons. Moreover, mapping to the Kyoto Encyclopedia of Genes and Genomes pathway database showed that the up-regulated proteins were mainly involved in metabolic pathways of tricarboxylic acid cycle, pyruvate metabolism, and nitrogen metabolism pathway, providing support for the metabolic balance of microbial cells shifting toward reduced end-products due to electron utilization. Using a biochemical method, the ompF-overexpressed strain was employed to investigate the function of the channel protein. These findings provided a theoretical basis for further improving electron transfer and utilization efficiency, and contributed to the potential applications of the bioelectrochemical system.</p

Image_1_Proteomics Reveal the Effect of Exogenous Electrons on Electroactive Escherichia coli.pdf

Microbial cells utilizing electricity to produce high-value fuels and chemicals are the foundation of the biocathodic bioelectrochemical system. However, molecular mechanisms of electron transfer and utilization have not been elucidated. In this work, Escherichia coli engineered by introducing the Mtr pathway from Shewanella oneidensis exhibited stronger electrochemical activity than control and could utilize exogenous electrons to stimulate metabolite profiles and boost succinate production in the bioelectrochemical system. Proteomic analysis and real-time PCR were performed to investigate the effect of exogenous electrons on electroactive E. coli. Bioinformatics analysis suggested that the proteins of molecular function associated with oxidoreductase activity, 4 iron, 4 sulfur([4Fe-4S]) cluster binding, iron-sulfur cluster binding, and metal cluster binding were positively affected by exogenous electrons. Moreover, mapping to the Kyoto Encyclopedia of Genes and Genomes pathway database showed that the up-regulated proteins were mainly involved in metabolic pathways of tricarboxylic acid cycle, pyruvate metabolism, and nitrogen metabolism pathway, providing support for the metabolic balance of microbial cells shifting toward reduced end-products due to electron utilization. Using a biochemical method, the ompF-overexpressed strain was employed to investigate the function of the channel protein. These findings provided a theoretical basis for further improving electron transfer and utilization efficiency, and contributed to the potential applications of the bioelectrochemical system.</p

Table_1_Evolutionary Diversity of Prophage DNA in Klebsiella pneumoniae Chromosomes.xlsx

Mobile gene elements play an important role in the continuous evolution of the prophage DNA of bacteria, promoting the emergence of new gene structures. This study explored the evolution of four strains of Klebsiella pneumoniae harboring prophages, 19051, 721005, 911021, and 675920, and 16 genomes of K. pneumoniae from GenBank. The results revealed a wide range of genetic variation in the prophage DNA inserted into the sap sites of K. pneumoniae chromosomes. From analysis and comparison of the sequences of the 20 prophage DNAs determined from the four strains and the 16 GenBank genomes of K. pneumoniae using high-throughput sequencing and antimicrobial susceptibility tests, we identified a novel transposon, Tn6556. We also identified at least nine large genetic structures with massive genetic acquisitions or losses and five hotspot sites showing a tendency to undergo insertion of gene elements such as IS1T, IS1R, IS26, ISKpn26, ISKpn28, Tn6556, MDR, and In27-related regions as variable regions; however, the only highly conserved core genes were int and umuCD among the 20 prophage DNAs. These findings provide important insights into the evolutionary diversity of bacteriophage DNA contained in K. pneumoniae.</p

Table_2_Evolutionary Diversity of Prophage DNA in Klebsiella pneumoniae Chromosomes.xlsx

Mobile gene elements play an important role in the continuous evolution of the prophage DNA of bacteria, promoting the emergence of new gene structures. This study explored the evolution of four strains of Klebsiella pneumoniae harboring prophages, 19051, 721005, 911021, and 675920, and 16 genomes of K. pneumoniae from GenBank. The results revealed a wide range of genetic variation in the prophage DNA inserted into the sap sites of K. pneumoniae chromosomes. From analysis and comparison of the sequences of the 20 prophage DNAs determined from the four strains and the 16 GenBank genomes of K. pneumoniae using high-throughput sequencing and antimicrobial susceptibility tests, we identified a novel transposon, Tn6556. We also identified at least nine large genetic structures with massive genetic acquisitions or losses and five hotspot sites showing a tendency to undergo insertion of gene elements such as IS1T, IS1R, IS26, ISKpn26, ISKpn28, Tn6556, MDR, and In27-related regions as variable regions; however, the only highly conserved core genes were int and umuCD among the 20 prophage DNAs. These findings provide important insights into the evolutionary diversity of bacteriophage DNA contained in K. pneumoniae.</p

Table_1_Engineering a Microbial Consortium Based Whole-Cell System for Efficient Production of Glutarate From L-Lysine.docx

Glutarate is an important C5 platform chemical produced during the catabolism of L-lysine through 5-aminovalerate (5-AMV) pathway. Here, we first established a whole-cell biocatalysis system for the glutarate production from L-lysine with the engineered Escherichia coli (E. coli) that co-expressed DavAB and GabDT. However, the accumulation of intermediate 5-AMV was identified as one important factor limiting glutarate production. Meanwhile, the negative interaction of co-expressing DavAB and GabDT in a single cell was also confirmed. Here, we solved these problems through engineering a microbial consortium composed of two engineered E. coli strains, BL21-22AB and BL21-YDT, as the whole-cell biocatalysts, each of which contains a part of the glutarate pathway. After the optimization of bioconversion conditions, including temperature, metal ion additives, pH, and cell ratio, 17.2 g/L glutarate was obtained from 20 g/L L-lysine with a yield of 95.1%, which was improved by 19.2% compared with that in a single cell. Little accumulation of 5-AMV was detected. Even at the high substrate concentration, the reduced 5-AMV accumulation and increased glutarate production were achieved. This synthetic consortium produced 43.8 g/L glutarate via a fed-batch strategy, the highest titer reported to date.</p