Accelerated Engineering of ELP-Based Materials through Hybrid Biomimetic-De Novo Predictive Molecular Design.

Efforts to engineer high-performance protein-based materials inspired by nature have mostly focused on altering naturally occurring sequences to confer the desired functionalities, whereas de novo design lags significantly behind and calls for unconventional innovative approaches. Here, using partially disordered elastin-like polypeptides (ELPs) as initial building blocks this work shows that de novo engineering of protein materials can be accelerated through hybrid biomimetic design, which this work achieves by integrating computational modeling, deep neural network, and recombinant DNA technology. This generalizable approach involves incorporating a series of de novo-designed sequences with α-helical conformation and genetically encoding them into biologically inspired intrinsically disordered repeating motifs. The new ELP variants maintain structural conformation and showed tunable supramolecular self-assembly out of thermal equilibrium with phase behavior in vitro. This work illustrates the effective translation of the predicted molecular designs in structural and functional materials. The proposed methodology can be applied to a broad range of partially disordered biomacromolecules and potentially pave the way toward the discovery of novel structural proteins.

Production of D-glucaric acid with phosphoglucose isomerase-deficient Saccharomyces cerevisiae.

D-Glucaric acid is a potential biobased platform chemical. Previously mainly Escherichia coli, but also the yeast Saccharomyces cerevisiae, and Pichia pastoris, have been engineered for conversion of D-glucose to D-glucaric acid via myo-inositol. One reason for low yields from the yeast strains is the strong flux towards glycolysis. Thus, to decrease the flux of D-glucose to biomass, and to increase D-glucaric acid yield, the four step D-glucaric acid pathway was introduced into a phosphoglucose isomerase deficient (Pgi1p-deficient) Saccharomyces cerevisiae strain. High D-glucose concentrations are toxic to the Pgi1p-deficient strains, so various feeding strategies and use of polymeric substrates were studied. Uniformly labelled 13C-glucose confirmed conversion of D-glucose to D-glucaric acid. In batch bioreactor cultures with pulsed D-fructose and ethanol provision 1.3 g D-glucaric acid L-1 was produced. The D-glucaric acid titer (0.71 g D-glucaric acid L-1) was lower in nitrogen limited conditions, but the yield, 0.23 g D-glucaric acid [g D-glucose consumed]-1, was among the highest that has so far been reported from yeast. Accumulation of myo-inositol indicated that myo-inositol oxygenase activity was limiting, and that there would be potential to even higher yield. The Pgi1p-deficiency in S. cerevisiae provides an approach that in combination with other reported modifications and bioprocess strategies would promote the development of high yield D-glucaric acid yeast strains.

Inducible Synthetic Growth Regulation Using the ClpXP Proteasome Enhances cis,cis-Muconic Acid and Glycolic Acid Yields in Saccharomyces cerevisiae.

Engineered microbial cells can produce sustainable chemistry, but the production competes for resources with growth. Inducible synthetic control over the resource use would enable fast accumulation of sufficient biomass and then divert the resources to production. We developed inducible synthetic resource-use control overSaccharomyces cerevisiae by expressing a bacterial ClpXP proteasome from an inducible promoter. By individually targeting growth-essential metabolic enzymes Aro1, Hom3, and Acc1 to the ClpXP proteasome, cell growth could be efficiently repressed during cultivation. The ClpXP proteasome was specific to the target proteins, and there was no reduction in the targets when ClpXP was not induced. The inducible growth repression improved product yields from glucose (cis,cis-muconic acid) and per biomass (cis,cis-muconic acid and glycolic acid). The inducible ClpXP proteasome tackles uncertainties in strain optimization by enabling model-guided repression of competing, growth-essential, and metabolic enzymes. Most importantly, it allows improving production without compromising biomass accumulation when uninduced; therefore, it is expected to mitigate strain stability and low productivity challenges.

The complex structure of Fomes fomentarius represents an architectural design for high-performance ultralightweight materials.

High strength, hardness, and fracture toughness are mechanical properties that are not commonly associated with the fleshy body of a fungus. Here, we show with detailed structural, chemical, and mechanical characterization that Fomes fomentarius is an exception, and its architectural design is a source of inspiration for an emerging class of ultralightweight high-performance materials. Our findings reveal that F. fomentarius is a functionally graded material with three distinct layers that undergo multiscale hierarchical self-assembly. Mycelium is the primary component in all layers. However, in each layer, mycelium exhibits a very distinct microstructure with unique preferential orientation, aspect ratio, density, and branch length. We also show that an extracellular matrix acts as a reinforcing adhesive that differs in each layer in terms of quantity, polymeric content, and interconnectivity. These findings demonstrate how the synergistic interplay of the aforementioned features results in distinct mechanical properties for each layer.

Correction: ß-1,3-Glucan synthesis, novel supramolecular self-assembly, characterization and application.

Correction for 'ß-1,3-Glucan synthesis, novel supramolecular self-assembly, characterization and application' by Robert Pylkkänen et al., Nanoscale, 2022, https://doi.org/10.1039/D2NR02731C.

ß-1,3-Glucan synthesis, novel supramolecular self-assembly, characterization and application.

ß-1,3-Glucans are ubiquitously observed in various biological systems with diverse physio-ecological functions, yet their underlying assembly mechanism and multiscale complexation in vitro remains poorly understood. Here, we provide for the first-time evidence of unidentified ß-1,3-glucan supramolecular complexation into intricate hierarchical architectures over several length scales. We mediated these unique assemblies using a recombinantly produced ß-1,3-glucan phosphorylase (Ta1,3BGP) by fine-tuning solution conditions during particle nucleation and growth. We report a synthesis of interconnected parallel hexagonal lamellae composed of 8 nm thick sheets of highly expanded paracrystals. The architecture consists of ß-1,3-glucan triple-helices with considerable inter-intra hydrogen bonding within, as well as in between adjacent triple-helices. The results extend our understanding of ß-1,3-glucan molecular organization and shed light on different aspects of the crystallization processes of biomolecules into structures unseen by nature. The presented versatile synthesis yields new materials for diverse medical and industrial applications.

Strain design optimization using reinforcement learning.

Engineered microbial cells present a sustainable alternative to fossil-based synthesis of chemicals and fuels. Cellular synthesis routes are readily assembled and introduced into microbial strains using state-of-the-art synthetic biology tools. However, the optimization of the strains required to reach industrially feasible production levels is far less efficient. It typically relies on trial-and-error leading into high uncertainty in total duration and cost. New techniques that can cope with the complexity and limited mechanistic knowledge of the cellular regulation are called for guiding the strain optimization. In this paper, we put forward a multi-agent reinforcement learning (MARL) approach that learns from experiments to tune the metabolic enzyme levels so that the production is improved. Our method is model-free and does not assume prior knowledge of the microbe's metabolic network or its regulation. The multi-agent approach is well-suited to make use of parallel experiments such as multi-well plates commonly used for screening microbial strains. We demonstrate the method's capabilities using the genome-scale kinetic model of Escherichia coli, k-ecoli457, as a surrogate for an in vivo cell behaviour in cultivation experiments. We investigate the method's performance relevant for practical applicability in strain engineering i.e. the speed of convergence towards the optimum response, noise tolerance, and the statistical stability of the solutions found. We further evaluate the proposed MARL approach in improving L-tryptophan production by yeast Saccharomyces cerevisiae, using publicly available experimental data on the performance of a combinatorial strain library. Overall, our results show that multi-agent reinforcement learning is a promising approach for guiding the strain optimization beyond mechanistic knowledge, with the goal of faster and more reliably obtaining industrially attractive production levels.

PHB production from cellobiose with Saccharomyces cerevisiae.

Replacement of petrochemical-based materials with microbially produced biodegradable alternatives calls for industrially attractive fermentation processes. Lignocellulosic materials offer non-edible alternatives for cultivated sugars, but require often use of expensive sugar releasing enzymes, such as ß-glucosidases. These cellulose treatment costs could be reduced if microbial production hosts could use short cellodextrins such as cellobiose directly as their substrates. In this study, we demonstrate production of poly(hydroxybutyrate) (PHB) in yeast Saccharomyces cerevisiae using cellobiose as a sole carbon source. Yeast strains expressing PHB pathway genes from Cupriavidus necator and cellodextrin transporter gene CDT-1 from Neurospora crassa were complemented either with ß-glucosidase gene GH1-1 from N. crassa or with cellobiose phosphorylase gene cbp from Ruminococcus flavefaciens. These cellobiose utilization routes either with Gh1-1 or Cbp enzymes differ in energetics and dynamics. However, both routes enabled higher PHB production per consumed sugar and higher PHB accumulation % of cell dry weight (CDW) than use of glucose as a carbon source. As expected, the strains with Gh1-1 consumed cellobiose faster than the strains with Cbp, both in flask and bioreactor batch cultures. In shake flasks, higher final PHB accumulation % of CDW was reached with Cbp route (10.0 ± 0.3%) than with Gh1-1 route (8.1 ± 0.2%). However, a higher PHB accumulation was achieved in better aerated and pH-controlled bioreactors, in comparison to shake flasks, and the relative performance of strains switched. In bioreactors, notable PHB accumulation levels per CDW of 13.4 ± 0.9% and 18.5 ± 3.9% were achieved with Cbp and Gh1-1 routes, respectively. The average molecular weights of accumulated PHB were similar using both routes; approximately 500 kDa and 450 kDa for strains expressing either cbp or GH1-1 genes, respectively. The formation of PHB with high molecular weights, combined with efficient cellobiose conversion, demonstrates a highly potential solution for improving attractiveness of sustainable polymer production using microbial cells.

Control of D-lactic acid content in P(LA-3HB) copolymer in the yeast Saccharomyces cerevisiae using a synthetic gene expression system.

The fully biobased polyhydroxyalkanoate (PHA) polymers provide interesting alternatives for petrochemical derived plastic materials. The mechanical properties of some PHAs, including the common poly(3-hydroxybutyrate) (PHB), are limited, but tunable by addition of other monomers into the polymer chain. In this study we present a precise synthetic biology method to adjust lactate monomer fraction of a polymer by controlling the monomer formation in vivo at gene expression level, independent of cultivation conditions. We used the modified doxycycline-based Tet-On approach to adjust the expression of the stereospecific D-lactate dehydrogenase gene (ldhA) from Leuconostoc mesenteroides to control D-lactic acid formation in yeast Saccharomyces cerevisiae. The synthetic Tet-On transcription factor with a VP16 activation domain was continuously expressed and its binding to a synthetic promoter with eight transcription factor specific binding sites upstream of the ldhA gene was controlled with the doxycycline concentration in the media. The increase in doxycycline concentration correlated positively with ldhA expression, D-lactic acid production, poly(D-lactic acid) (PDLA) accumulation in vivo, and D-lactic acid content in the poly(D-lactate-co-3-hydroxybutyrate) P(LA-3HB) copolymer. We demonstrated that the D-lactic acid content of the P(LA-3HB) copolymer can be adjusted linearly from 6 mol% to 93 mol% in vivo in S. cerevisiae. These results highlight the power of controlling gene expression and monomer formation in the tuning of the polymer composition. In addition, we obtained 5.6% PDLA and 19% P(LA-3HB) of the cell dry weight (CDW), which are over two- and five-fold higher accumulation levels, respectively, than reported in the previous studies with yeast. We also compared two engineered PHA synthases and discovered that in S. cerevisiae the PHA synthase PhaC1437Ps6-19 produced P(LA-3HB) copolymers with lower D-lactic acid content, but with higher molecular weight, in comparison to the PHA synthase PhaC1Pre.

Bioinspired Functionally Graded Composite Assembled Using Cellulose Nanocrystals and Genetically Engineered Proteins with Controlled Biomineralization.

Nature provides unique insights into design strategies evolved by living organisms to construct robust materials with a combination of mechanical properties that are challenging to replicate synthetically. Hereby, inspired by the impact-resistant dactyl club of the stomatopod, a mineralized biocomposite is rationally designed and produced in the complex shapes of dental implant crowns exhibiting high strength, stiffness, and fracture toughness. This material consists of an expanded helicoidal organization of cellulose nanocrystals (CNCs) mixed with genetically engineered proteins that regulate both binding to CNCs and in situ growth of reinforcing apatite crystals. Critically, the structural properties emerge from controlled self-assembly across multiple length scales regulated by rational engineering and phase separation of the protein components. This work replicates multiscale biomanufacturing of a model biological material and also offers an innovative platform to synthesize multifunctional biocomposites whose properties can be finely regulated by colloidal self-assembly and engineering of its constitutive protein building blocks.

Production of D-lactic acid containing polyhydroxyalkanoate polymers in yeast Saccharomyces cerevisiae.

Polyhydroxyalkanoates (PHAs) provide biodegradable and bio-based alternatives to conventional plastics. Incorporation of 2-hydroxy acid monomers into polymer, in addition to 3-hydroxy acids, offers possibility to tailor the polymer properties. In this study, poly(D-lactic acid) (PDLA) and copolymer P(LA-3HB) were produced and characterized for the first time in the yeast Saccharomyces cerevisiae. Expression of engineered PHA synthase PhaC1437Ps6-19, propionyl-CoA transferase Pct540Cp, acetyl-CoA acetyltransferase PhaA, and acetoacetyl-CoA reductase PhaB1 resulted in accumulation of 3.6% P(LA-3HB) and expression of engineered enzymes PhaC1Pre and PctMe resulted in accumulation of 0.73% PDLA of the cell dry weight (CDW). According to NMR, P(LA-3HB) contained D-lactic acid repeating sequences. For reference, expression of PhaA, PhaB1, and PHA synthase PhaC1 resulted in accumulation 11% poly(hydroxybutyrate) (PHB) of the CDW. Weight average molecular weights of these polymers were comparable to similar polymers produced by bacterial strains, 24.6, 6.3, and 1 130 kDa for P(LA-3HB), PDLA, and PHB, respectively. The results suggest that yeast, as a robust and acid tolerant industrial production organism, could be suitable for production of 2-hydroxy acid containing PHAs from sugars or from 2-hydroxy acid containing raw materials. Moreover, the wide substrate specificity of PHA synthase enzymes employed increases the possibilities for modifying copolymer properties in yeast in the future.

Bioprocess performance analysis of novel methanol-independent promoters for recombinant protein production with Pichia pastoris.

BACKGROUND: Pichia pastoris is a powerful and broadly used host for recombinant protein production (RPP), where past bioprocess performance has often been directed with the methanol regulated AOX1 promoter (PAOX1), and the constitutive GAP promoter (PGAP). Since promoters play a crucial role in an expression system and the bioprocess efficiency, innovative alternatives are constantly developed and implemented. Here, a thorough comparative kinetic characterization of two expression systems based on the commercial PDF and UPP promoters (PPDF, PUPP) was first conducted in chemostat cultures. Most promising conditions were subsequently tested in fed-batch cultivations. These new alternatives were compared with the classical strong promoter PGAP, using the Candida antarctica lipase B (CalB) as model protein for expression system performance. RESULTS: Both the PPDF and PUPP-based expression systems outperformed similar PGAP-based expression in chemostat cultivations, reaching ninefold higher specific production rates (qp). CALB transcription levels were drastically higher when employing the novel expression systems. This higher expression was also correlated with a marked upregulation of unfolded protein response (UPR) related genes, likely from an increased protein burden in the endoplasmic reticulum (ER). Based on the chemostat results obtained, best culture strategies for both PPDF and PUPP expression systems were also successfully implemented in 15 L fed-batch cultivations where qp and product to biomass yield (YP/X*) values were similar than those obtained in chemostat cultivations. CONCLUSIONS: As an outcome of the macrokinetic characterization presented, the novel PPDF and PUPP were observed to offer much higher efficiency for CalB production than the widely used PGAP-based methanol-free alternative. Thus, both systems arise as highly productive alternatives for P. pastoris-based RPP bioprocesses. Furthermore, the different expression regulation patterns observed indicate the level of gene expression can be adjusted, or tuned, which is interesting when using Pichia pastoris as a cell factory for different products of interest.

NordAqua, a Nordic Center of Excellence to develop an algae-based photosynthetic production platform.

NordAqua is a multidisciplinary Nordic Center of Excellence funded by NordForsk Bioeconomy program (2017-2022). The research center promotes Blue Bioeconomy and endeavours to reform the use of natural resources in a environmentally sustainable way. In this short communication, we summarize particular outcomes of the consortium. The key research progress of NordAqua includes (1) improving of photosynthetisis, (2) developing novel photosynthetic cell factories that function in a "solar-driven direct CO2 capture to target bioproducts" mode, (3) promoting the diversity of Nordic cyanobacteria and algae as an abundant and resilient alternative for less sustainable forest biomass and for innovative production of biochemicals, and (4) improving the bio-based wastewater purification and nutrient recycling technologies to provide new tools for integrative circular economy platforms.

Substrate specificity of 2-deoxy-D-ribose 5-phosphate aldolase (DERA) assessed by different protein engineering and machine learning methods.

In this work, deoxyribose-5-phosphate aldolase (Ec DERA, EC 4.1.2.4) from Escherichia coli was chosen as the protein engineering target for improving the substrate preference towards smaller, non-phosphorylated aldehyde donor substrates, in particular towards acetaldehyde. The initial broad set of mutations was directed to 24 amino acid positions in the active site or in the close vicinity, based on the 3D complex structure of the E. coli DERA wild-type aldolase. The specific activity of the DERA variants containing one to three amino acid mutations was characterised using three different substrates. A novel machine learning (ML) model utilising Gaussian processes and feature learning was applied for the 3rd mutagenesis round to predict new beneficial mutant combinations. This led to the most clear-cut (two- to threefold) improvement in acetaldehyde (C2) addition capability with the concomitant abolishment of the activity towards the natural donor molecule glyceraldehyde-3-phosphate (C3P) as well as the non-phosphorylated equivalent (C3). The Ec DERA variants were also tested on aldol reaction utilising formaldehyde (C1) as the donor. Ec DERA wild-type was shown to be able to carry out this reaction, and furthermore, some of the improved variants on acetaldehyde addition reaction turned out to have also improved activity on formaldehyde. KEY POINTS: • DERA aldolases are promiscuous enzymes. • Synthetic utility of DERA aldolase was improved by protein engineering approaches. • Machine learning methods aid the protein engineering of DERA.

In Vitro Synthesis and Self-Assembly of Cellulose II Nanofibrils Catalyzed by the Reverse Reaction of Clostridium thermocellum Cellodextrin Phosphorylase.

In nature, various organisms produce cellulose as microfibrils, which are processed into their nano- and microfibrillar and/or crystalline components by humans in order to obtain desired material properties. Interestingly, the natural synthesis machinery can be circumvented by enzymatically synthesizing cellulose from precursor molecules in vitro. This approach is appealing for producing tailor-made cellulosic particles and materials because it enables optimization of the reaction conditions for cellulose synthesis in order to generate particles with a desired morphology in their pure form. Here, we present enzymatic cellulose synthesis catalyzed by the reverse reaction of Clostridium thermocellum cellodextrin phosphorylase in vitro. We were able to produce cellulose II nanofibril networks in all conditions tested, using varying concentrations of the glycosyl acceptors d-glucose or d-cellobiose (0.5, 5, and 50 mM). We show that shorter cellulose chains assemble into flat ribbon-like fibrils with greater diameter, while longer chains assemble into cylindrical fibrils with smaller diameter.

The Lipomyces starkeyi gene Ls120451 encodes a cellobiose transporter that enables cellobiose fermentation in Saccharomyces cerevisiae.

Processed lignocellulosic biomass is a source of mixed sugars that can be used for microbial fermentation into fuels or higher value products, like chemicals. Previously, the yeast Saccharomyces cerevisiae was engineered to utilize its cellodextrins through the heterologous expression of sugar transporters together with an intracellular expressed ß-glucosidase. In this study, we screened a selection of eight (putative) cellodextrin transporters from different yeast and fungal hosts in order to extend the catalogue of available cellobiose transporters for cellobiose fermentation in S. cerevisiae. We confirmed that several in silico predicted cellodextrin transporters from Aspergillus niger were capable of transporting cellobiose with low affinity. In addition, we found a novel cellobiose transporter from the yeast Lipomyces starkeyi, encoded by the gene Ls120451. This transporter allowed efficient growth on cellobiose, while it also grew on glucose and lactose, but not cellotriose nor cellotetraose. We characterized the transporter more in-depth together with the transporter CdtG from Penicillium oxalicum. CdtG showed to be slightly more efficient in cellobiose consumption than Ls120451 at concentrations below 1.0 g/L. Ls120451 was more efficient in cellobiose consumption at higher concentrations and strains expressing this transporter grew slightly slower, but produced up to 30% more ethanol than CdtG.

Solar-Powered Carbon Fixation for Food and Feed Production Using Microorganisms-A Comparative Techno-Economic Analysis.

This study evaluates the techno-economic feasibility of five solar-powered concepts for the production of autotrophic microorganisms for food and feed production; the main focus is on three concepts based on hydrogen-oxidizing bacteria (HOB), which are further compared to two microalgae-related concepts. Two locations with markedly different solar conditions are considered (Finland and Morocco), in which Morocco was found to be the most economically competitive for the cultivation of microalgae in open ponds and closed systems (1.4 and 1.9 € kg-1, respectively). Biomass production by combined water electrolysis and HOB cultivation results in higher costs for all three considered concepts. Among these, the lowest production cost of 5.3 € kg-1 is associated with grid-assisted electricity use in Finland, while the highest production cost of >9.1 € kg-1 is determined for concepts using solely photovoltaics and/or photoelectrochemical technology for on-site electricity production and solar-energy conversion to H2 by water electrolysis. All assessed concepts are capital intensive. Furthermore, a sensitivity analysis suggests that the production costs of HOB biomass can be lowered down to 2.1 € kg-1 by optimization of the process parameters among which volumetric productivity, electricity strategy, and electricity costs have the highest cost-saving potentials. The study reveals that continuously available electricity and H2 supply are essential for the development of a viable HOB concept due to the capital intensity of the needed technologies. In addition, volumetric productivity is the key parameter that needs to be optimized to increase the economic competitiveness of HOB production.

Controllable coacervation of recombinantly produced spider silk protein using kosmotropic salts.

Recent developments suggest that the phase transition of natural and synthetic biomacromolecules represents an important and ubiquitous mechanism underlying structural assemblies toward the fabrication of high-performance materials. Such a transition results in the formation of condensed liquid droplets, described as condensates or coacervates. Being able to effectively control the assembly of such entities is essential for tuning the quality and their functionality. Here we describe how self-coacervation of genetically engineered spidroin-inspired proteins can be preceded by a wide range of kosmotropic salts. We studied the kinetics and mechanisms of coacervation in different conditions, from direct observation of initial phase separation to the early stage of nucleation/growth and fusion into large fluid assemblies. We found that coacervation induced by kosmotropic salts follows the classical nucleation theory and critically relies on precursor clusters of few weak-interacting protein monomers. Depending on solution conditions and the strength of the supramolecular interaction as a function of time, coacervates with a continuum of physiochemical properties were observed. We observed similar characteristics in other protein-based coacervates, which include having a spherical-ellipsoid shape in solution, an interconnected bicontinuous network, surface adhesion, and wetting properties. Finally, we demonstrated the use of salt-induced self-coacervates of spidroin-inspired protein as a cellulosic binder in dried condition.

Evaluation of synthetic formaldehyde and methanol assimilation pathways in Yarrowia lipolytica.

BACKGROUND: Crude glycerol coming from biodiesel production is an attractive carbon source for biological production of chemicals. The major impurity in preparations of crude glycerol is methanol, which is toxic for most microbes. Development of microbes, which would not only tolerate the methanol, but also use it as co-substrate, would increase the feasibility of bioprocesses using crude glycerol as substrate. RESULTS: To prevent methanol conversion to CO2 via formaldehyde and formate, the formaldehyde dehydrogenase (FLD) gene was identified in and deleted from Yarrowia lipolytica. The deletion strain was able to convert methanol to formaldehyde without expression of heterologous methanol dehydrogenases. Further, it was shown that expression of heterologous formaldehyde assimilating enzymes could complement the deletion of FLD. The expression of either 3-hexulose-6-phosphate synthase (HPS) enzyme of ribulose monosphosphate pathway or dihydroxyacetone synthase (DHAS) enzyme of xylulose monosphosphate pathway restored the formaldehyde tolerance of the formaldehyde sensitive Δfld1 strain. CONCLUSIONS: In silico, the expression of heterologous formaldehyde assimilation pathways enable Y. lipolytica to use methanol as substrate for growth and metabolite production. In vivo, methanol was shown to be converted to formaldehyde and the enzymes of formaldehyde assimilation were actively expressed in this yeast. However, further development is required to enable Y. lipolytica to efficiently use methanol as co-substrate with glycerol.

Towards patterned bioelectronics: facilitated immobilization of exoelectrogenic Escherichia coli with heterologous pili.

Biosensors detect signals using biological sensing components such as redox enzymes and biological cells. Although cellular versatility can be beneficial for different applications, limited stability and efficiency in signal transduction at electrode surfaces represent a challenge. Recent studies have shown that the Mtr electron conduit from Shewanella oneidensis MR-1 can be produced in Escherichia coli to generate an exoelectrogenic model system with well-characterized genetic tools. However, means to specifically immobilize this organism at solid substrates as electroactive biofilms have not been tested previously. Here, we show that mannose-binding Fim pili can be produced in exoelectrogenic E. coli and can be used to selectively attach cells to a mannose-coated material. Importantly, cells expressing fim genes retained current production by the heterologous Mtr electron conduit. Our results demonstrate the versatility of the exoelectrogenic E. coli system and motivate future work that aims to produce patterned biofilms for bioelectronic devices that can respond to various biochemical signals.