Enhancing Power Generation In Shewanella By Regulation Of Flavins Metabolism And Biofilm Modification

Shewanella is a group of microorganisms that have the ability of extracellular electron transfer(EET),which breaks down organic matter and converts stored chemical energy into electricity.However,due to the low concentration of flavins and poor conductivity of biofilm,the power generation of Shewanella is weak,which limits its application in industry.In this paper,EET efficiency was improved from three aspects:increasing the yield of flavins,enhancing the concentration of c-Cyts and improving the formation of biofilm.The main research results are as follows:1.In vitro addition of flavins to construct microbial fuel cells(MFCs)In vitro adding of different concentrations and proportions of flavins construct MFCs to explore which flavin is the most important electron shuttle of the EET process.The results showed that the highest power was 2350±119 m W/m~2 in the MFC containing 40μmol/L riboflavin.These results indicated that riboflavin is the main extracellular electron shuttle compared with flavin mononucleotide(FMN)and flavin adenine dinucleotide(FAD).2.Construction of riboflavin synthetic strains guided by in vitro flavin addition experiments Using promoter and different copy number plasmid strategies,riboflavin operons from Bacillus subtilis and Escherichia coli were heterologously introduced to construct strains with high levels of flavin.The results showed that under the induction of high copy number plasmid PHG13 and medium intensity promoter Ptet,the strain Ptet-5 overexpressing the riboflavin operon of E.coli reached the highest flavin yield of 889.3μmol/L.The power density of 707±95 m W/m~2 was 16.44 times that of WT(MR-1)(10.5μmol/L).3.Construction of strains overexpressing c-type cytochrome protein Overexpressing omc A,mtr C,mtr A,mtr B,cym A,and mtr ABC,respectively in Ptet5 analyze which protein interacts with RF in MFCs.The results showed that under the induction of 0.5mmol/L IPTG,the strain Ptet-5/PL-Mtr C overexpression showed the best performance,and the power density reached 1046±87 m W/m~2,which was 36%higher than that of the control strain.This indicates that the riboflavin mainly interacts with Mtr C in MFCs.4.Construction strains via the modulation of cell morphology to promote the biofilm formation Overexpressing of sul A,min CD,mre B and ftsz,respectively in Ptet-5 screen genes that regulate biofilm formation.The results showed that the power density of Ptet-5/Psul A with overexpression of sul A was greatly improved,reaching 1414±19m W/m~2,which was twice that of the control strain.This indicates that the gene sul A regulating cell enlargement can enhance the formation of biofilm.5.Construction of multi-module modified strains of flavin,cytochrome protein and cell morphology Ptet-5/PMtr C-sul A was obtained by module assembly of E.coli riboflavin operon,cytochrome protein Mtr C and gene sul A encoding inhibitory cell division protein by multi-module combination strategy.The results showed that the electric power density of the engineered strain was increased by 56%(1104±59 m W/m~2)compared with the control strain under 3 mmol/L arabinose induction.