An Escherichia coli Chassis for Production of Electrically Conductive Protein Nanowires

Abstract

11Geobacter sulfurreducens' pilin-based electrically conductive protein nanowires (e-PNs) are a 12 revolutionary electronic material. They offer novel options for electronic sensing applications 13 and have the remarkable ability to harvest electrical energy from atmospheric humidity. 14 However, technical constraints limit mass cultivation and genetic manipulation of G. 15 sulfurreducens. Therefore, we designed a strain of Escherichia coli to express e-PNs by 16 introducing a plasmid that contained an inducible operon with E. coli genes for type IV pili 17 biogenesis machinery and a synthetic gene designed to yield a peptide monomer that could be 18 assembled into e-PNs. The e-PNs expressed in E. coli, and harvested with a simple filtration 19 method, had the same diameter (3 nm) and conductance as e-PNs expressed in G. 20 sulfurreducens. These results, coupled with the robustness of E. coli for mass cultivation and the 21 extensive E. coli toolbox for genetic manipulation, greatly expands opportunities for large-scale 22 fabrication of novel e-PNs. 23 24

Microbial biofilms for electricity generation from water evaporation and power to wearables

“…4b), or 43% and 19% values of the same size biofilm-sheet devices. Devices fabricated with mats of genetically modified E. coli expressing conductive protein nanowires 32 , generated an average voltage of 0.25 V and a current of 0.18 µA (Fig. 4b), or 54% and 31% values of the same size biofilm-sheet devices.…”

Engineering Wired Life: Synthetic Biology for Electroactive Bacteria

“…First, a partly refactored pili assembly machinery from enterohemorrhagic E. coli 103 was expressed on a plasmid with a substituted pili monomer (pilA). 104 The conductance of resulting pilin was similar to that of those extracted from G. sulfurreducens. Second, a scalable approach to producing pilin monomers from recombinant PilA shows that they selfassemble into protein nanowires with properties consistent with G. sulf urreducens.…”

“…Second, a scalable approach to producing pilin monomers from recombinant PilA shows that they selfassemble into protein nanowires with properties consistent with G. sulf urreducens. 43,104 This work lays a foundation for precise biosynthesis of pili nanowires, which can be further harnessed to derive functionalized versions and will doubtless stimulate exciting applications in bioelectrical devices 105,106 and bioinspired synthetic materials. 107 ■ ENGINEERED BIOELECTRONIC SIGNALING WITH SOLUBLE REDOX ACTIVE MOLECULES As described above, soluble redox active mediators play a well established role in microorganisms with EET capabilities.…”

Biomaterials and Electroactive Bacteria for Biodegradable Electronics

“…The introduction of the PilA gene from Geobacter metallireducens in G. sulfurreducens , e.g., produces e-pili with conductivities in the order of 300 S cm –1 ( Tan et al, 2017 ), while the inclusion of more aromatic rings like tryptophan in e-pili increased the conductivities to 100 S cm –1 at pH 7 and 1,000 S cm –1 at pH 2 ( Tan et al, 2016 ). When thinking about applying these bacterial nanowires in biodegradable electronics, one might assume an intensive and costly process, but the recent discovery of nanowire production through genetic manipulation of Escherichia coli ( Ueki et al, 2020 ) and the development of bottom-up fabrication of e-pili ( Cosert et al, 2019 ) suggest a means of easier and cheaper production in the long term. Functionalization and adhesion of different substrates is also made easier with the possibility of decorating nanowires with peptides ( Ueki et al, 2019 ).…”

“…In the search for biological materials that could function as conductors or transistors in biodegradable electronics, interesting candidates are found in the world of electromicrobiology ( Lovley, 2012 ), where electroactive bacteria like Geobacter and Shewanella species produce nanowires as electron transport conduits. Recent progress in the study of different proteins ( Yalcin et al, 2020 ) and genetic manipulation has ramped up the conductivity values ( Tan et al, 2016 , 2017 ); modification of Escherichia coli has made PilA based nanowires easy to produce ( Ueki et al, 2020 ). Yet, conduction lengths exceeding micrometers are yet to be shown, as well as the implementation in biodegradable electronic circuitry.…”