Cytochromes C Research Articles

Single-Molecule Detection of the Encounter and Productive Electron Transfer Complexes of a Photosynthetic Reaction Center.

Small, diffusible redox proteins play an essential role in electron transfer (ET) in respiration and photosynthesis, sustaining life on Earth by shuttling electrons between membrane-bound complexes via finely tuned and reversible interactions. Ensemble kinetic studies show transient ET complexes form in two distinct stages: an "encounter" complex largely mediated by electrostatic interactions, which subsequently, through subtle reorganization of the binding interface, forms a "productive" ET complex stabilized by additional hydrophobic interactions around the redox-active cofactors. Here, using single-molecule force spectroscopy (SMFS) we dissected the transient ET complexes formed between the photosynthetic reaction center-light harvesting complex 1 (RC-LH1) of Rhodobacter sphaeroides and its native electron donor cytochrome c2 (cyt c2). Importantly, SMFS resolves the distribution of interaction forces into low (∼150 pN) and high (∼330 pN) components, with the former more susceptible to salt concentration and to alteration of key charged residues on the RC. Thus, the low force component is suggested to reflect the contribution of electrostatic interactions in forming the initial encounter complex, whereas the high force component reflects the additional stabilization provided by hydrophobic interactions to the productive ET complex. Employing molecular dynamics simulations, we resolve five intermediate states that comprise the encounter, productive ET and leaving complexes, predicting a weak interaction between cyt c2 and the LH1 ring near the RC-L subunit that could lie along the exit path for oxidized cyt c2. The multimodal nature of the interactions of ET complexes captured here may have wider implications for ET in all domains of life.

Effect of pH on the thermostability and redox properties of cytochrome c552 from Wolinella succinogenes

Effect of pH on the thermostability and redox properties of cytochrome c552 from Wolinella succinogenes

APX2 Is an Ascorbate Peroxidase-Related Protein that Regulates the Levels of Plastocyanin in Chlamydomonas.

The function of ascorbate peroxidase-related (APX-R) proteins, present in all green photosynthetic eukaryotes, remains unclear. This study focuses on APX-R from Chlamydomonas reinhardtii, namely, ascorbate peroxidase 2 (APX2). We showed that apx2 mutants exhibited a faster oxidation of the photosystem I primary electron donor, P700, upon sudden light increase and a slower re-reduction rate compared to the wild type, pointing to a limitation of plastocyanin. Spectroscopic, proteomic and immunoblot analyses confirmed that the phenotype was a result of lower levels of plastocyanin in the apx2 mutants. The redox state of P700 did not differ between wild type and apx2 mutants when the loss of function in plastocyanin was nutritionally complemented by growing apx2 mutants under copper deficiency. In this case, cytochrome c6 functionally replaces plastocyanin, confirming that lower levels of plastocyanin were the primary defect caused by the absence of APX2. Overall, the results presented here shed light on an unexpected regulation of plastocyanin level under copper-replete conditions, induced by APX2 in Chlamydomonas.

Antimalarial Potential of Phytochemical Compounds from Garcinia atroviridis Griff ex. T. Anders Targeting Multiple Proteins of Plasmodium falciparum 3D7: An In Silico Approach

Abstract Background: Plasmodium falciparum is a malaria-causing unicellular parasite with strain 3D7 (Pf3D7) being the most lethal. Currently, antimalarial resistance has been reported which necessitates the development of novel antimalarial drugs to combat the spread of malaria. Garcinia atroviridis Griff. ex T. Anders contains phytochemical compounds that are useful for various activities, including targeting Pf3D7 proteins. This study explored novel antimalarial drugs from G. atroviridis against several target proteins of Pf3D7 in silico. Methods: Phytocompounds from G. atroviridis were selected as ligands. After retrieval from the Protein Data Bank, the protein sequence was screened using BLASTp NCBI. Molecular docking analysis was performed on PyRx to compute binding affinity and identify the chemical interactions involved. The stability of the ligand-protein complex was evaluated using dynamic molecular approaches. Results: Our findings showed that quercetin has a high binding affinity with apicoplast DNA polymerase (−8.3 kcal/mol), glutamyl-tRNA synthetase (−7.5 kcal/mol), and plasmepsin X (−7.8 kcal/mol). Kaempferol had a high binding affinity for the cytochrome c2 domain-swapped dimer (−8.4 kcal/mol). Conclusion: Collectively, quercetin and kaempferol are potential antimalarial candidates which warrant further investigation using in vitro and in vivo designs.

Roadmap of electrons from donor side to the reaction center of photosynthetic purple bacteria with mutated cytochromes

In photosynthetic bacteria, the absorbed light drives the canonical cyclic electron transfer between the reaction center and the cytochrome bc1 complexes via the pools of mobile electron carriers. If kinetic or structural barriers hinder the participation of the bc1 complex in the cyclic flow of electrons, then the pools of mobile redox agents must supply the electrons for the multiple turnovers of the reaction center. These conditions were achieved by continuous high light excitation of intact cells of bacterial strains Rba. sphaeroides and Rvx. gelatinosus with depleted donor side cytochromes c2 (cycA) and tetraheme cytochrome subunit (pufC), respectively. The gradual oxidation by ferricyanide further reduced the availability of electron donors to pufC. Electron transfer through the reaction center was tracked by absorption change and by induction and relaxation of the fluorescence of the bacteriochlorophyll dimer. The rate constants of the electron transfer (~ 3 × 103 s‒1) from the mobile donors of Rvx. gelatinosus bound either to the RC (pufC) or to the tetraheme subunit (wild type) were similar. The electrons transferred through the reaction center dimer were supplied entirely by the donor pool; their number amounted to about 5 in wild type Rvx. gelatinosus and decreased to 1 in pufC oxidized by ferricyanide. Fluorescence yield was measured as a function of the oxidized fraction of the dimer and its complex shape reveals the contribution of two competing processes: the migration of the excitation energy among the photosynthetic units and the availability of electron donors to the oxidized dimer. The experimental results were simulated and rationalized by a simple kinetic model of the two-electron cycling of the acceptor side combined with aperiodic one-electron redox function of the donor side.

Juglone, a plant-derived 1,4-naphthoquinone, binds to hydroxylamine oxidoreductase and inhibits the electron transfer to cytochrome c554.

Nitrification, the microbial conversion of ammonia to nitrate via nitrite, plays a pivotal role in the global nitrogen cycle. However, the excessive use of ammonium-based fertilizers in agriculture has disrupted this cycle, leading to groundwater pollution and greenhouse gas emissions. In this study, we have demonstrated the inhibitory effects of plant-derived juglone and related 1,4-naphthoquinones on the nitrification process in Nitrosomonas europaea. Notably, the inhibition mechanism is elucidated in which 1,4-naphthoquinones interact with hydroxylamine oxidoreductase, disrupting the electron transfer to cytochrome c554, a physiological electron acceptor. These findings support the notion that phytochemicals can impede nitrification by interfering with the essential electron transfer process in ammonia oxidation. The findings presented in this article offer valuable insights for the development of strategies aimed at the management of nitrification, reduction of fertilizer utilization, and mitigation of greenhouse gas emissions.

An engineered thermally tolerant apo-cytochrome scaffold for metal-less incorporation of heme derivative.

Cytochrome c552 from Thermus thermophilus is one of the hot topics for creating smart biomaterials as it possesses remarkable stability, is tolerant to multiple mutations and has therefore been recently reported for a number of functionalizations upon substitution of the original prosthetic group with an artificial prosthetic group. However, all of the substitutions were driven by the coordination through the axial ligands followed by complete reconstitution with a metal-porphyrin complex. This limits the scope of the cytochrome c for incorporating a metal-less non-natural heme species that could improve the versatility of cytochrome c for a new generation of engineered cytochrome proteins for further enhancement in their functionalities such as biocatalysts. In this connection, a new variant of Cytochrome c (rC552 C14A) from Thermus thermophilus was reported, where an easy approach to remove the original prosthetic group was achieved, followed by the incorporation of a number of metal-PPIX derivatives that ultimately led to the formation of artificial c-type cytochromes through covalent bonding. The apo-cytochrome was found to be thermally tolerant and to possess a distinctive overall structure as that of the wild type, as was evident from the corresponding CD spectra, which ultimately encouraged reconstitution with a metal-less protoporphyrin derivative for better understanding the role of axial ligands in the reconstitution process. Successful reconstitution was achieved, resulting in a new type of Cytochrome b-type artificial protein without the metal in its active site, indicating the non-involvement of the axial ligand. In order to prove the non-involvement of the axial ligand, a subsequent double mutant (C14A/M69A) was constructed, replacing the methionine at 69 position with non-coordinating alanine residue. Accordingly, the apo-C14A/M69A was prepared and found to be extremely stable as the earlier mutants and the WT showed no signs of denaturation, even at the elevated temperature of 98°C. Subsequently, heme b was successfully incorporated into the apo-C14A/M69A, which demonstrated itself as a highly thermally tolerant protein scaffold for incorporating a metal-less artificial prosthetic group in the absence of the axial ligand. Further improvement in the reconstitution process is achieved by replacing the methionine at 69 position with phenyl alanine (C14A/M69F mutant), resulting in further stabilization of heme species, possibly through non-covalent π-interactions, as corroborated by molecular docking.

Methacrylate Redox Systems of Anaerobic Bacteria

The review analyzes current information about the anaerobic type of respiration using a non-natural methacrylate compound as an electron acceptor. Both the methacrylate redox systems themselves and the anaerobic bacteria in whose cells they are found are considered. These complexes consist of flavin-containing reductase and multiheme cytochrome(s) c3. The genes of the components of the methacrylate redox systems of different microorganisms are homologous and are organized into one operon. Methacrylate-reducing activity is determined in the periplasm. The only known bacterial acrylate reductase that reduces the natural compound differs from methacrylate redox systems. The physiological role, origin, and research perspectives for this unique enzyme system are discussed.

Structure of the bc1–cbb3 respiratory supercomplex from Pseudomonas aeruginosa

Energy conversion by electron transport chains occurs through the sequential transfer of electrons between protein complexes and intermediate electron carriers, creating the proton motive force that enables ATP synthesis and membrane transport. These protein complexes can also form higher order assemblies known as respiratory supercomplexes (SCs). The electron transport chain of the opportunistic pathogen Pseudomonas aeruginosa is closely linked with its ability to invade host tissue, tolerate harsh conditions, and resist antibiotics but is poorly characterized. Here, we determine the structure of a P. aeruginosa SC that forms between the quinol:cytochrome c oxidoreductase (cytochrome bc1) and one of the organism's terminal oxidases, cytochrome cbb3, which is found only in some bacteria. Remarkably, the SC structure also includes two intermediate electron carriers: a diheme cytochrome c4 and a single heme cytochrome c5. Together, these proteins allow electron transfer from ubiquinol in cytochrome bc1 to oxygen in cytochrome cbb3. We also present evidence that different isoforms of cytochrome cbb3 can participate in formation of this SC without changing the overall SC architecture. Incorporating these different subunit isoforms into the SC would allow the bacterium to adapt to different environmental conditions. Bioinformatic analysis focusing on structural motifs in the SC suggests that cytochrome bc1-cbb3 SCs also exist in other bacterial pathogens.

A cytochrome c551 mediates the cyclic electron transport chain of the anoxygenic phototrophic bacterium Roseiflexus castenholzii

Roseiflexus castenholzii is a gram-negative filamentous phototrophic bacterium that carries out anoxygenic photosynthesis through a cyclic electron transport chain (ETC). The ETC is composed of a reaction center (RC)–light-harvesting (LH) complex (rcRC–LH); an alternative complex III (rcACIII), which functionally replaces the cytochrome bc1/b6f complex; and the periplasmic electron acceptor auracyanin (rcAc). Although compositionally and structurally different from the bc1/b6f complex, rcACIII plays similar essential roles in oxidizing menaquinol and transferring electrons to the rcAc. However, rcACIII-mediated electron transfer (which includes both an intraprotein route and a downstream route) has not been clearly elucidated, nor have the details of cyclic ETC. Here, we identify a previously unknown monoheme cytochrome c (cyt c551) as a novel periplasmic electron acceptor of rcACIII. It reduces the light-excited rcRC–LH to complete a cyclic ETC. We also reveal the molecular mechanisms involved in the ETC using electron paramagnetic resonance (EPR), spectroelectrochemistry, and enzymatic and structural analyses. We find that electrons released from rcACIII-oxidized menaquinol are transferred to two alternative periplasmic electron acceptors (rcAc and cyt c551), which eventually reduce the rcRC to form the complete cyclic ETC. This work serves as a foundation for further studies of ACIII-mediated electron transfer in anoxygenic photosynthesis and broadens our understanding of the diversity and molecular evolution of prokaryotic ETCs.

A Comparative Multi-Frequency EPR Study of Dipolar Interaction in Tetra-Heme Cytochromes.

Distances between Fe ions in multiheme cytochromes are sufficiently short to make the intramolecular dipole-dipole interaction between hemes probable. In the analysis of EPR data from cytochromes, this interaction has thus far been ignored under the assumption that spectra are the simple sum of non-interacting components. Here, we use a recently developed low-frequency broadband EPR spectrometer to establish the extent of dipolar interaction in the example cytochromes, characterize its spectral signatures, and identify present limitations in the analysis. Broadband EPR spectra of Shewanella oneidensis MR-1 small tetraheme cytochrome (STC) have been collected over the frequency range of 0.45 to 13.11 GHz, and they have been compared to similar data from Desulfovibrio vulgaris Hildenborough cytochrome c3. The two cases are representative examples of two very different heme topologies and corresponding electron-transfer properties in tetraheme proteins. While in cytochrome c3, the six Fe-Fe distances can be sorted into two well-separated groups, those in STC are diffuse. Since the onset of dipolar interaction between Fe-Fe pairs is already observed in the X-band, the g values are determined in the simulation of the 13.11 GHz spectrum. Low-frequency spectra are analyzed with the inclusion of dipolar interaction based on available structural data on mutual distances and orientations between all hemes. In this procedure, all 24 possible assignments of individual heme spectra to heme topologies are sampled. The 24 configurations can be reduced to a few, but inspection falls short of a unique assignment, due to a remaining lack of understanding of the fine details of these complex spectra. In general, the EPR analysis suggests the four-heme system in c3 to be more rigid than that in STC, which is proposed to be related to different physiological roles in electron transfer.

Ecofriendly Approach on the Removal of Reactive Orange 107 from Aqueous Solutions Using Cladophora Species as a Novel Biosorbent.

The efficiency of Cladophora species for the removal of Reactive Orange 107 (RO107) from the aqueous solution was evaluated through batch adsorption studies by optimising various process parameters such as pH (3-8), dye concentration (100-500mg/l), biosorbent concentration (100-500mg/l), temperature (25-45°C) and contact time (12-108h). The results revealed that the optimum conditions for RO107 decolourisation (87%) was found on 72h of incubation with 100mg/l dye concentration amended with 200mg/l biosorbent at pH 6 at 25°C. The mechanism of dye adsorption was evaluated using isotherms, kinetics and thermodynamic models. The experimental data fitted well with Langmuir isotherm and pseudo-second-order kinetic models. Thermodynamic studies revealed that the adsorption process was endothermic, spontaneous and feasible in nature. Recovery of RO107 from the Cladophora sp. was maximum when 0.1M HNO3 was used as an eluent. UV-Visible, FT-IR and SEM analyses reveal the interaction between the biosorbent-adsorbate and confirm the process of decolourisation by Cladophora sp. In order to evaluate the nature of the untreated and treated dye solutions, toxicological studies were conducted and the results revealed that the treated dye solution was non- toxic as compared with untreated dye solution. The results of the docking study proved that there was a substantial binding energy between RO107 and the protein (Cytochrome C6) of Cladophora sp. Hence, Cladophora sp. proves to be a promising biosorbent to decolourise RO107 and its potential can be explored in the textile sectors.

The structure of the complex between the arsenite oxidase from Pseudorhizobium banfieldiae sp. strain NT-26 and its native electron acceptor cytochrome c552.

The arsenite oxidase (AioAB) from Pseudorhizobium banfieldiae sp. strain NT-26 catalyzes the oxidation of arsenite to arsenate and transfers electrons to its cognate electron acceptor cytochrome c552 (cytc552). This activity underpins the ability of this organism to respire using arsenite present in contaminated environments. The crystal structure of the AioAB/cytc552 electron transfer complex reveals two A2B2/(cytc552)2 assemblies per asymmetric unit. Three of the four cytc552 molecules in the asymmetric unit dock to AioAB in a cleft at the interface between the AioA and AioB subunits, with an edge-to-edge distance of 7.5 Å between the heme of cytc552 and the [2Fe-2S] Rieske cluster in the AioB subunit. The interface between the AioAB and cytc552 proteins features electrostatic and nonpolar interactions and is stabilized by two salt bridges. A modest number of hydrogen bonds, salt bridges and relatively small, buried surface areas between protein partners are typical features of transient electron transfer complexes. Interestingly, the fourth cytc552 molecule is positioned differently between two AioAB heterodimers, with distances between its heme and the AioAB redox active cofactors that are outside the acceptable range for fast electron transfer. This unique cytc552 molecule appears to be positioned to facilitate crystal packing rather than reflecting a functional complex.

Toxicity Effects of Perfluorooctanoic Acid (PFOA) and Perfluorooctane Sulfonate (PFOS) on Two Green Microalgae Species.

Amongst per- and polyfluoroalkyl substances (PFAS) compounds, perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS) have a high persistence in physicochemical and biological degradation; therefore, the accumulation of PFOS and PFOA can negatively affect aquatic organisms and human health. In this study, two microalgae species (Chlorella vulgaris and Scenedesmus obliquus) were exposed to different concentrations of a PFOS and PFOA mixture (0 to 10 mg L-1). With increases in the contact time (days) and the PFAS concentration (mg L-1) from 1 to 7, and 0.5 to 10, respectively, the cell viability, total chlorophyll content, and protein content decreased, and the decrease in these parameters was significantly greater in Scenedesmus obliquus. As another step in the study, the response surface methodology (RSM) was used to optimize the toxicity effects of PFAS on microalgae in a logical way, as demonstrated by the high R2 (>0.9). In another stage, a molecular docking study was performed to monitor the interaction of PFOS and PFOA with the microalgae, considering hydrolysis and the enzymes involved in oxidation-reduction reactions using individual enzymes. The analysis was conducted on carboxypeptidase in Chlorella vulgaris and on c-terminal processing protease and oxidized cytochrome c6 in Scenedesmus obliquus. For the enzyme activity, the affinity and dimensions of ligands-binding sites and ligand-binding energy were estimated in each case.

Soluble domains of cytochrome c-556 and Rieske iron-sulfur protein from Chlorobaculum tepidum: Crystal structures and interaction analysis.

In photosynthetic green sulfur bacteria, the electron transfer reaction from menaquinol:cytochrome c oxidoreductase to the P840 reaction center (RC) complex occurs directly without any involvement of soluble electron carrier protein(s). X-ray crystallography has determined the three-dimensional structures of the soluble domains of the CT0073 gene product and Rieske iron-sulfur protein (ISP). The former is a mono-heme cytochrome c with an α-absorption peak at 556nm. The overall fold of the soluble domain of cytochrome c-556 (designated as cyt c-556sol) consists of four α-helices and is very similar to that of water-soluble cyt c-554 that independently functions as an electron donor to the P840 RC complex. However, the latter's remarkably long and flexible loop between the α3 and α4 helices seems to make it impossible to be a substitute for the former. The structure of the soluble domain of the Rieske ISP (Rieskesol protein) shows a typical β-sheets-dominated fold with a small cluster-binding and a large subdomain. The architecture of the Rieskesol protein is bilobal and belongs to those of b6f-type Rieske ISPs. Nuclear magnetic resonance (NMR) measurements revealed weak non-polar but specific interaction sites on Rieskesol protein when mixed with cyt c-556sol. Therefore, menaquinol:cytochrome c oxidoreductase in green sulfur bacteria features a Rieske/cytb complex tightly associated with membrane-anchored cyt c-556.

Drop-casted Photosystem I/cytochrome c multilayer films for biohybrid solar energy conversion.

One of the main barriers to making efficient Photosystem I-based biohybrid solar cells is the need for an electrochemical pathway to facilitate electron transfer between the P700 reaction center of Photosystem I and an electrode. To this end, nature provides inspiration in the form of cytochrome c6, a natural electron donor to the P700 site. Its natural ability to access the P700 binding pocket and reduce the reaction center can be mimicked by employing cytochrome c, which has a similar protein structure and redox chemistry while also being compatible with a variety of electrode surfaces. Previous research has incorporated cytochrome c to improve the photocurrent generation of Photosystem I using time consuming and/or specialized electrode preparation. While those methods lead to high protein areal density, in this work we use the quick and facile vacuum-assisted drop-casting technique to construct a Photosystem I/cytochrome c photoactive composite film with micron-scale thickness. We demonstrate that this simple fabrication technique can result in high cytochrome c loading and improvement in cathodic photocurrent over a drop-casted Photosystem I film without cytochrome c. In addition, we analyze the behavior of the cytochrome c/Photosystem I system at varying applied potentials to show that the improvement in performance can be attributed to enhancement of the electron transfer rate to P700 sites and therefore the PSI turnover rate within the composite film.

Separation and analysis of Bacillus subtilis respiratory chain complexes.

Bacillus subtilis is a Gram-positive bacterium with a respiratory chain embedded in the cytoplasmic membrane. The respiratory chain is bifurcated after menaquinol into a cytochrome b6c + caa3 branch and a branch with up to three quinol oxidases. The complexes that generate the proton gradient are b6c, associated with caa3 and aa3 oxidase. The b6c and caa3 complexes form a supercomplex, and it is proposed to form respiratory strings in the membrane. There is still information missing about the quinol branch and if the primary oxidase quinol aa3 is associated with the electron donor complexes. It is unclear whether succinate quinone reductase (SQR) can form associations with the quinol branch or the cytochrome branch. In this paper, we show the separation of an almost pure b6c complex associated with cytochromes c550 and c551. We obtained a b6c + caa3 supercomplex of 600kDa and SQR, aa3, and NADH dehydrogenase by dodecyl maltoside solubilization and separation of the respiratory chain components by ionic exchange chromatography. We found that aa3 does not associate with other complexes. SQR was associated with the b6c complex in a mutant lacking aa3. This association could facilitate electron transfer from SQR to menaquinone-7. The lack of associations between the abundant quinol oxidase aa3 and other complexes is a feature we cannot explain yet.

Perchlorate reduction kinetics and genome-resolved metagenomics identify metabolic interactions in acclimated saline lake perchlorate-reducing consortia

Perchlorate is a widely detected environmental contaminant in surface and underground water, that seriously impacts human health by inhibiting the uptake of thyroidal radioiodine. Perchlorate reduction due to saline lake microorganisms is not as well understood as that in marine environments. In this study, we enriched a perchlorate-reducing microbial consortium collected from saline lake sediments and found that the perchlorate reduction kinetics of the enriched consortium fit the Michaelis–Menten kinetics well, with a maximum specific substrate reduction rate (qmax) of 0.596 ± 0.001 mg ClO4−/mg DW/h and half-saturation constant (Ks) of 16.549 ± 0.488 mg ClO4−/L. Furthermore, we used improved metagenome binning to reconstruct high-quality metagenome-assembled genomes from the metagenomes of the microbial consortia, including the perchlorate-reducing bacteria (PRB) Dechloromonas agitata and Wolinella succinogenes, with the genome of W. succinogenes harboring complete functional genes for perchlorate reduction being the first recovered. Given that the electrons were directly transferred to the electronic carrier cytochrome c-553 from the quinone pool, the electron transfer pathway of W. succinogenes was shorter and more efficient than the canonical pattern. This finding provides a theoretical basis for microbial remediation of sites contaminated by high concentrations of perchlorate. Metagenomic binning and metatranscriptomic analyses revealed the gene transcription variation of perchlorate reductase pcr and chlorite dismutase cld by PRB and the synergistic metabolic mechanism.

The preparation of PANI-doped Co-Ni-ZIF carbonized derivatives and the exploration of EET process by the DFT calculation and the prediction of related functional genes

As a green conversion/energy storage device, the practical application of microbial fuel cell (MFC) was limited with its low power generation. The extracellular electron transfer efficiency was one of the key factors affecting the power generation performance of MFC. This paper reported the carbonized derivative Co-Ni-ZIF@NC derived from PANI-doped Co-Ni-ZIF with conductivity, biocompatibility and high electrocatalytic activity. Density functional theory (DFT) confirmed the strong interaction between the electrode surface and microorganisms. The maximum power density of the MFC with Co-Ni-ZIF@NC anode (8.67 W/m3) was 1.47 time higher than that of Co-Ni-ZIF@C-800 (5.96 W/m3). High-throughput sequencing revealed that Co-Ni-ZIF@NC catalyst facilitated the screening of relevant functional microbial communities in microbial membranes, leading to the reinforcement of power generation in MFC. PICRUSt predicted the relative abundance of functional genes, confirmed the spontaneous enrichment behaviour of functional microorganisms to the electrode surface, and identified cytochrome c552 as the dominant role in the EET process. The above foregoing mainly due to the doping of PANI, improved the content of pyrrolic N on the surface of derivatives, protected the original crystal morphology, promoted the approaching behaviour of conductive flagella to the anode surface and shortened the electron transport distance, thus Co-Ni-ZIF@NC exhibited superb bioelectrocatalytic activity.

Manganese oxidation counteracts the deleterious effect of low temperatures on biofilm formation in Pseudomonas sp. MOB-449.

Mn removal from groundwater by biological sand filter technology is negatively impacted by low temperatures in winter periods. Therefore, the need to study Mn(II)-oxidizing bacteria (MOB) having the potential to oxidize Mn(II) and form biofilms at low temperatures is imperative. These MOB can have potential as inocula for sand filter bioaugmentation strategies to optimize Mn removal during winter periods. We previously showed that a Pseudomonas sp. MOB-449 (MOB-449), isolated from a Mn biofilter, oxidizes Mn(II) in a biofilm-dependent way at low temperatures. In this work, MOB-449 Mn(II) oxidation and growth capacities were evaluated under planktonic and biofilm conditions at different temperatures. At 18°C, MOB-449 showed enhanced biofilm formation due to the addition of Mn(II) to the medium correlating with Mn(II) oxidation, compared to biofilms grown in control medium. Moreover, this enhancement on biofilm formation due to the addition of Mn(II) was only observed at 18°C. At this temperature, Mn(II) oxidation in membrane fractions collected from biofilms was induced by uncoupling oxidative phosphorylation from the electron transport chain with 2,4-Dinitrophenol. In Pseudomonas, a role for c-type cytochrome in Mn(II) oxidation has been demonstrated. Accordingly, transcriptional profiles of all terminal oxidases genes found in MOB-449 showed an induction of cytochrome c terminal oxidases expression mediated by Mn(II) oxidation at 18°C. Finally, heme peroxidase activity assays and MS analysis revealed that PetC, a cytochrome c5, and also CcmE, involved in the cytochrome c biogenesis machinery, are induced at 18°C only in the presence of Mn(II). These results present evidence supporting that cytochromes c and also the cytochrome c terminal oxidases are activated at low temperatures in the presence of Mn(II). Overall, this work demonstrate that in MOB-449 Mn(II) oxidation is activated at low temperatures to gain energy, suggesting that this process is important for survival under adverse environmental conditions and contributing to the understanding of the physiological role of bacterial Mn(II) oxidation.