The discovery of lytic polysaccharide monooxygenases (LPMOs), a family of copper-dependent enzymes that play a major role in polysaccharide degradation, has revealed the importance of oxidoreductases in the biological utilization of biomass. In fungi, a range of redox proteins have been implicated as working in harness with LPMOs to bring about polysaccharide oxidation. In bacteria, less is known about the interplay between redox proteins and LPMOs, or how the interaction between the two contributes to polysaccharide degradation. We therefore set out to characterize two previously unstudied proteins from the shipworm symbiont Teredinibacter turnerae that were initially identified by the presence of carbohydrate binding domains appended to uncharacterized domains with probable redox functions. Here, X-ray crystal structures of several domains from these proteins are presented together with initial efforts to characterize their functions. The analysis suggests that the target proteins are unlikely to function as LPMO electron donors, raising new questions as to the potential redox functions that these large extracellular multi-haem-containing c-type cytochromes may perform in these bacteria.

The interaction of bacteria and archaea with electrodes is a relatively new research field which spans from fundamental to applied research and influences interdisciplinary research in the fields of microbiology, biochemistry, biotechnology as well as process engineering. Although a substantial understanding of electron transfer processes between microbes and anodes and between microbes and cathodes has been achieved in mesophilic organisms, the mechanisms used by microbes under extremophilic conditions are still in the early stages of discovery. Here, we review our current knowledge on the biochemical solutions that evolved for the interaction of extremophilic organisms with electrodes. To this end, the available knowledge on pure cultures of extremophilic microorganisms has been compiled and the study has been extended with the help of bioinformatic analyses on the potential distribution of different electron transfer mechanisms in extremophilic microorganisms.

Sporomusa ovata is a bacterium that can accept electrons from cathodes to drive microbial electrosynthesis (MES) of acetate from carbon dioxide. It is the biocatalyst with the highest acetate production rate described. Here we review the research on S. ovata across different disciplines, including microbiology, biochemistry, engineering, and materials science, to summarize and assess the state-of-the-art. The improvement of the biocatalytic capacity of S. ovata in the last 10 years, using different optimization strategies is described and discussed. In addition, we propose possible electron uptake routes derived from genetic and experimental data described in the literature and point out the possibilities to understand and improve the performance of S. ovata through genetic engineering. Finally, we identify current knowledge gaps guiding further research efforts to explore this promising organism for the MES field.

Direct interspecies electron transfer (DIET) is an effective mechanism for microbial species to exchange electrons cooperatively during syntrophic metabolism. It is generally accepted that DIET is mainly mediated by electrically conductive pili and outer surface c-type cytochromes (c-Cyts). However, as an extracellular matrix is ubiquitous and abundant on the surface of microorganisms, the effect and mechanism of exopolysaccharides on DIET are still unclear. This study constructed a co-culture of exopolysaccharides-deficient Geobacter sulfurreducens with Geobacter metallireducens to explore the role of exopolysaccharides in DIET. Results revealed that the deficiency of exopolysaccharides extended the metabolic period of the co-culture by 44.4% and changed the proportions of each species in the co-culture. The exopolysaccharides-deficient co-culture failed to form large, tight spherical aggregates and the expression of c-Cyts and pili was decreased. The addition of magnetite and granular activated carbon (GAC), respectively, might compensate for the functions of c-Cyts and pili in the first generation of co-culture, but the stimulatory effect on the metabolic stable period co-culture was fairly limited. These findings demonstrate that non-conductive exopolysaccharides are an important component of DIET aggregates and an extracellular matrix for DIET-required c-Cyts.

Uranium (U) pollution is an environmental hazard caused by the development of the nuclear industry. Microbial reduction of hexavalent uranium (U(VI)) to tetravalent uranium (U(IV)) reduces U solubility and mobility and has been proposed as an effective method to remediate uranium contamination. In this review, U(VI) remediation with respect to U(VI)-reducing bacteria, mechanisms, influencing factors, products, and reoxidation are systematically summarized. Reportedly, some metal- and sulfate-reducing bacteria possess excellent U(VI) reduction capability through mechanisms involving c-type cytochromes, extracellular pili, electron shuttle, or thioredoxin reduction. In situ remediation has been demonstrated as an ideal strategy for large-scale degradation of uranium contaminants than ex situ. However, U(VI) reduction efficiency can be affected by various factors, including pH, temperature, bicarbonate, electron donors, and coexisting metal ions. Furthermore, it is noteworthy that the reduction products could be reoxidized when exposed to oxygen and nitrate, inevitably compromising the remediation effects, especially for non-crystalline U(IV) with weak stability.

Electroactive biofilm (EAB) has been considered as the core determining electricity generation in microbial fuel cells (MFCs), and its spatial structure regulation for enhanced activity and selectivity is of great concern. In this study, iron phthalocyanine (FePc) was introduced into a carbon cloth (CC) electrode, aiming at improving the affinity between the anode and outer membrane c-type cytochromes (OM c-Cyts) and achieving a highly active EAB. The FePc modified CC anode (FePc-CC) effectively improved the viability of EAB and enriched the Geobacter species up to 44.83% (FePc-CC) from 6.97% (CC). The FePc-CC anode achieved a much higher power density of 2419 mW m-2 than the CC (560 mW m-2) and a remarkable higher biomass loading of 2477.2 ± 84.5 μg cm-2 than the CC (749.3 ± 31.3 μg cm-2). As the charge transfer resistance was decreased by 58.6 times from 395.2 Ω (CC) to 6.74 Ω (FePc-CC), the interfacial reaction rate was accelerated and the direct electron transfer via OM c-Cyts was promoted. This work provides an effective method to improve the EAB activity by regulating its spatial structure, and opens the door toward the development of highly active EAB using metal phthalocyanines in MFCs.

Geobacter sulfurreducens biofilms have promising applications in renewable energy, pollutant bioremediation, and bioelectronic applications. Genetically manipulating G. sulfurreducens biofilms is an effective strategy to improve the capacity of extracellular electron transfer (EET). Extracellular polysaccharide, a sticky component surrounding microbes, plays an important role in EET. Herein, we constructed a mutant of G. sulfurreducens strain PCA overexpressing the gene GSU1501 (part of the ATP-dependent exporter of the polysaccharide biosynthesis gene operon), designated strain PCA-1501, to increase EET capacity. Experimental results showed that the overexpression of GSU1501 increased extracellular polysaccharide secretion by 25.5%, which promoted the formation of biofilm with higher thickness and viability, as well as the content of extracellular c-type cytochromes. Compared with the control strain, the mutant showed a higher capacity of Fe(III) oxide reduction and current generation (increased by 20.4% and 22.2%, respectively). Interestingly, the overexpression of GSU1501 hindered the pili formation by reducing the transcription level of pilA; a compensatory relationship between extracellular polysaccharide and pili in promoting biofilm formation deserves further investigation. This study provides a feasible method to promote the EET capacity of G. sulfurreducens biofilms, which benefit their bioelectrochemical applications.

A shift from petrochemical processes toward a bio-based economy is one of the most advocated developments for a sustainable future. To achieve this will require the biotechnological production of platform chemicals that can be further processed by chemical engineering. Bioelectrochemical systems (BESs) are a novel tool within the biotechnology field. In BESs, microbes serve as biocatalysts for the production of biofuels and value-added compounds, as well as for the production of electricity. Although the general feasibility of bioelectrochemical processes has been demonstrated in recent years, much research has been conducted to develop biocatalysts better suited to meet industrial demands. Initially, mainly natural exoelectrogenic organisms were investigated for their performance in BESs. Driven by possibilities of recent developments in genetic engineering and synthetic biology, the spectrum of microbial catalysts and their versatility (substrate and product range) have expanded significantly. Despite these developments, there is still a tremendous gap between currently achievable space-time yields and current densities on the one hand and the theoretical limits of BESs on the other. It will be necessary to move the performance of the biocatalysts closer to the theoretical possibilities in order to establish viable production routines. This review summarizes the status quo of engineering microbial biocatalysts for anode-applications with high space-time yields. Furthermore, we will address some of the theoretical limitations of these processes exemplarily and discuss which of the present strategies might be combined to achieve highly synergistic effects and, thus, meet industrial demands.

Artemisinin and its derivatives kill malaria parasites and inhibit the proliferation of cancer cells. In both processes, heme was shown to play a key role in artemisinin bioactivation. We found that artemisinin and clinical artemisinin derivatives are able to compensate for a mutation in the yeast Bcs1 protein, a key chaperon involved in biogenesis of the mitochondrial respiratory complex III. The equivalent Bcs1 variant causes an encephalopathy in human by affecting complex III assembly. We show that artemisinin derivatives decrease the content of mitochondrial cytochromes and disturb the maturation of the complex III cytochrome c1. This last effect is likely responsible for the compensation by decreasing the detrimental over-accumulation of the inactive pre-complex III observed in the bcs1 mutant. We further show that a fluorescent dihydroartemisinin probe rapidly accumulates in the mitochondrial network and targets cytochromes c and c1 in yeast, human cells and isolated mitochondria. In vitro this probe interacts with purified cytochrome c only under reducing conditions and we detect cytochrome c-dihydroartemisinin covalent adducts by mass spectrometry analyses. We propose that reduced mitochondrial c-type cytochromes act as both targets and mediators of artemisinin bioactivation in yeast and human cells.

Graphene-based materials have been demonstrated to facilitate electron extracellular transfer (EET) of Shewanella. In this study, compared to group lacking graphene oxide (GO)-based materials, GO films-added group and graphene oxide/polyvinyl alcohol (GO/PVA) film-added group delivered 2.67- and 3.13-fold increases in the Cr(VI) reduction by Shewanella xiamenensis, respectively. The whole reduction process could be divided into three stages, including microbial Cr(VI) reduction and GO reduction stage, microbial GO reduction stage and microbial Cr(VI) reduction mediated by reduced graphene oxide (rGO) stage. Moreover, gene analysis revealed that addition of GO and GO/PVA films stimulated overexpression of several c-type cytochrome (c-Cyts) genes, including mtrA, mtrB, mtrC, mtrD, mtrE, mtrF, omcA, petC and SO-4047. Specifically, appreciable Cr(VI) reduction by the strains that overexpressed mtrA, mtrB, mtrC, mtrD, mtrE, mtrF and omcA further confirmed that overexpression of c-Cyts genes indeed enhanced the efficiency of Cr(VI) reduction. Based on these results, the specific function of every c-Cyt was clearly found in Cr(VI) reduction by the induction of GO-based materials. Our finding has disclosed a synergetic mechanism stimulated by GO-based materials to enhance Cr(VI) bioreduction that was not only mediated through the modification of material but also upregulated the expression of functional genes.