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1.
J Cell Sci ; 134(9)2021 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-34550353

RESUMO

Cytochrome c6 is a redox carrier in the thylakoid lumen of cyanobacteria and some eukaryotic algae. Although the isofunctional plastocyanin is present in land plants and the green alga Chlamydomonas reinhardtii, these organisms also possess a cytochrome c6-like protein designated as cytochrome c6A. Two other cytochrome c6-like groups, c6B and c6C, have been identified in cyanobacteria. In this study, we have identified a novel c6-like cytochrome called PetJ2, which is encoded in the nuclear genome of Cyanophora paradoxa, a member of the glaucophytes - the basal branch of the Archaeplastida. We propose that glaucophyte PetJ2 protein is related to cyanobacterial c6B and c6C cytochromes, and that cryptic green algal and land plant cytochromes c6A evolved from an ancestral archaeplastidial PetJ2 protein. In vitro import experiments with isolated muroplasts revealed that PetJ2 is imported into plastids. Although it harbors a twin-arginine motif in its thylakoid-targeting peptide, which is generally indicative of thylakoid import via the Tat import pathway, our import experiments with isolated muroplasts and the heterologous pea thylakoid import system revealed that PetJ2 uses the Sec pathway instead of the Tat import pathway.


Assuntos
Cyanophora , Sequência de Aminoácidos , Cyanophora/metabolismo , Citocromos/metabolismo , Eucariotos/metabolismo , Plastídeos/metabolismo
2.
Proc Natl Acad Sci U S A ; 118(39)2021 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-34556577

RESUMO

Proteins achieve efficient energy storage and conversion through electron transfer along a series of redox cofactors. Multiheme cytochromes are notable examples. These proteins transfer electrons over distance scales of several nanometers to >10 µm and in so doing they couple cellular metabolism with extracellular redox partners including electrodes. Here, we report pump-probe spectroscopy that provides a direct measure of the intrinsic rates of heme-heme electron transfer in this fascinating class of proteins. Our study took advantage of a spectrally unique His/Met-ligated heme introduced at a defined site within the decaheme extracellular MtrC protein of Shewanella oneidensis We observed rates of heme-to-heme electron transfer on the order of 109 s-1 (3.7 to 4.3 Å edge-to-edge distance), in good agreement with predictions based on density functional and molecular dynamics calculations. These rates are among the highest reported for ground-state electron transfer in biology. Yet, some fall 2 to 3 orders of magnitude below the Moser-Dutton ruler because electron transfer at these short distances is through space and therefore associated with a higher tunneling barrier than the through-protein tunneling scenario that is usual at longer distances. Moreover, we show that the His/Met-ligated heme creates an electron sink that stabilizes the charge separated state on the 100-µs time scale. This feature could be exploited in future designs of multiheme cytochromes as components of versatile photosynthetic biohybrid assemblies.


Assuntos
Grupo dos Citocromos c/metabolismo , Citocromos/metabolismo , Elétrons , Heme/metabolismo , Histidina/metabolismo , Metionina/metabolismo , Shewanella/metabolismo , Grupo dos Citocromos c/química , Citocromos/química , Transporte de Elétrons , Heme/química , Histidina/química , Metionina/química , Simulação de Dinâmica Molecular , Nanofios , Oxirredução
3.
PLoS One ; 16(8): e0244260, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34424897

RESUMO

Mitochondrial alternative oxidase (AOX) is predicted to be present in mitochondria of several invertebrate taxa including tardigrades. Independently of the reason concerning the enzyme occurrence in animal mitochondria, expression of AOX in human mitochondria is regarded as a potential therapeutic strategy. Till now, relevant data were obtained due to heterologous AOX expression in cells and animals without natively expressed AOX. Application of animals natively expressing AOX could importantly contribute to the research. Thus, we decided to investigate AOX activity in intact specimens of the tardigrade Hypsibius exemplaris. We observed that H. exemplaris specimens' tolerance to the blockage of the mitochondrial respiratory chain (MRC) cytochrome pathway was diminished in the presence of AOX inhibitor and the inhibitor-sensitive respiration enabled the tardigrade respiration under condition of the blockage. Importantly, these observations correlated with relevant changes of the mitochondrial inner membrane potential (Δψ) detected in intact animals. Moreover, detection of AOX at protein level required the MRC cytochrome pathway blockage. Overall, we demonstrated that AOX activity in tardigrades can be monitored by the animals' behavior observation as well as by measurement of intact specimens' whole-body respiration and Δψ. Furthermore, it is also possible to check the impact of the MRC cytochrome pathway blockage on AOX level as well as AOX inhibition in the absence of the blockage on animal functioning. Thus, H. exemplaris could be consider as a whole-animal model suitable to study AOX.


Assuntos
Mitocôndrias/metabolismo , Mitocôndrias/fisiologia , Proteínas Mitocondriais/metabolismo , Oxirredutases/metabolismo , Proteínas de Plantas/metabolismo , Tardígrados/metabolismo , Tardígrados/fisiologia , Animais , Comportamento Animal/fisiologia , Respiração Celular/fisiologia , Citocromos/metabolismo , Potencial da Membrana Mitocondrial/fisiologia , Membranas Mitocondriais/metabolismo , Membranas Mitocondriais/fisiologia , Oxirredução , Transdução de Sinais/fisiologia
4.
Photosynth Res ; 149(3): 265-273, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34228227

RESUMO

Martin Kamen was a giant of twentieth century science. Trained as a physical chemist, he was the co-discoverer of radioactive Carbon 14, which has transformed many areas of science as a tracer and as a way to date artifacts. He later switched to the study of metabolism and biochemistry and made important contributions to the understanding of nitrogen fixation and photosynthesis. Finally, he studied cytochromes, primarily from anoxygenic photosynthetic bacteria.


Assuntos
Bactérias/metabolismo , Carbono/química , Citocromos/química , Citocromos/metabolismo , Fotossíntese/fisiologia , História do Século XX , História do Século XXI , Humanos , Masculino , Estados Unidos
5.
Environ Sci Technol ; 55(14): 10142-10151, 2021 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-34196176

RESUMO

Direct interspecies electron transfer (DIET) between microbial species prevails in some key microbial consortia. However, the electron transfer mechanism(s) in these consortia is controversial due to lack of efficient characterization methods. Here, we provide an in situ anaerobic spectroelectrochemical coculture cell (in situ ASCC) to induce the formation of DIET coculture biofilm on the interdigitated microelectrode arrays and characterize the electron transfer directly. Two typical Geobacter DIET cocultures, Geobacter metallireducens and wild-type Geobacter sulfurreducens (G.m&G.s) and G. metallireducens and a G. sulfurreducens strain deficient in citrate synthase (G.m&G.s-ΔgltA), were selected. In situ Raman and electrochemical Fourier transform infrared (FTIR) spectroscopy indicated that cytochromes are abundant in the electric syntrophic coculture. Cyclic voltammetry and potential step experiment revealed a diffusion-controlled electron transfer process and the electrochemical gating measurements further demonstrated a cytochrome-mediated electron transfer in the DIET coculture. Furthermore, the G.m&G.s-ΔgltA coculture displayed a higher redox conductivity than the G.m&G.s coculture, consistent with the existence of an intimate and efficient electrical connection between these two species. Our findings provide the first report of a redox-gradient-driven electron transport facilitated by c-type cytochromes in DIET coculture, supporting the model that DIET is mediated by cytochromes and suggest a platform to explore the other DIET consortia.


Assuntos
Geobacter , Técnicas de Cocultura , Citocromos/metabolismo , Transporte de Elétrons , Geobacter/metabolismo , Oxirredução
6.
Sci Rep ; 11(1): 15140, 2021 07 23.
Artigo em Inglês | MEDLINE | ID: mdl-34302023

RESUMO

Interspecies hydrogen transfer (IHT) and direct interspecies electron transfer (DIET) are two syntrophy models for methanogenesis. Their relative importance in methanogenic environments is still unclear. Our recent discovery of a novel species Candidatus Geobacter eutrophica with the genetic potential of IHT and DIET may serve as a model species to address this knowledge gap. To experimentally demonstrate its DIET ability, we performed electrochemical enrichment of Ca. G. eutrophica-dominating communities under 0 and 0.4 V vs. Ag/AgCl based on the presumption that DIET and extracellular electron transfer (EET) share similar metabolic pathways. After three batches of enrichment, Geobacter OTU650, which was phylogenetically close to Ca. G. eutrophica, was outcompeted in the control but remained abundant and active under electrochemical stimulation, indicating Ca. G. eutrophica's EET ability. The high-quality draft genome further showed high phylogenomic similarity with Ca. G. eutrophica, and the genes encoding outer membrane cytochromes and enzymes for hydrogen metabolism were actively expressed. A Bayesian network was trained with the genes encoding enzymes for alcohol metabolism, hydrogen metabolism, EET, and methanogenesis from dominant fermentative bacteria, Geobacter, and Methanobacterium. Methane production could not be accurately predicted when the genes for IHT were in silico knocked out, inferring its more important role in methanogenesis. The genomics-enabled machine learning modeling approach can provide predictive insights into the importance of IHT and DIET.


Assuntos
Transporte de Elétrons/fisiologia , Geobacter/metabolismo , Hidrogênio/metabolismo , Teorema de Bayes , Citocromos/metabolismo , Elétrons , Aprendizado de Máquina , Redes e Vias Metabólicas/fisiologia , Metano/metabolismo , Methanobacterium/metabolismo
7.
Photosynth Res ; 148(3): 137-152, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34236566

RESUMO

The heliobacteria, a family of anoxygenic phototrophs, possess the simplest known photosynthetic apparatus. Although they are photoheterotrophs in the light, the heliobacteria can also grow chemotrophically via pyruvate metabolism in the dark. In the heliobacteria, the cytochrome bc complex is responsible for oxidizing menaquinol and reducing cytochrome c553 in the electron flow cycle used for phototrophy. However, there is no known electron acceptor for the mobile cytochrome c553 other than the photochemical reaction center. We have, therefore, hypothesized that the cytochrome bc complex is necessary for phototrophy, but unnecessary for chemotrophic growth in the dark. We used a two-step method for CRISPR-based genome editing in Heliobacterium modesticaldum to delete the genes encoding the four major subunits of the cytochrome bc complex. Genotypic analysis verified the deletion of the petCBDA gene cluster encoding the catalytic components of the complex. Spectroscopic studies revealed that re-reduction of cytochrome c553 after flash-induced photo-oxidation was over 100 times slower in the ∆petCBDA mutant compared to the wild-type. Steady-state levels of oxidized P800 (the primary donor of the photochemical reaction center) were much higher in the ∆petCBDA mutant at every light level, consistent with a limitation in electron flow to the reaction center. The ∆petCBDA mutant was unable to grow phototrophically on acetate plus CO2 but could grow chemotrophically on pyruvate as a carbon source similar to the wild-type strain in the dark. The mutants could be complemented by reintroduction of the petCBDA gene cluster on a plasmid expressed from the clostridial eno promoter.


Assuntos
Sobrevivência Celular/fisiologia , Clostridiales/genética , Clostridiales/metabolismo , Citocromos/genética , Citocromos/metabolismo , Deleção de Genes , Fotossíntese/fisiologia , Adaptação Ocular/genética , Adaptação Ocular/fisiologia , Adaptação à Escuridão/genética , Adaptação à Escuridão/fisiologia , Mutação , Fotossíntese/genética
8.
Biochem J ; 478(14): 2871-2887, 2021 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-34190983

RESUMO

The redox potential values of cytochromes can be modulated by the protonation/deprotonation of neighbor groups (redox-Bohr effect), a mechanism that permits the proteins to couple electron/proton transfer. In the respiratory chains, this effect is particularly relevant if observed in the physiological pH range, as it may contribute to the electrochemical gradient for ATP synthesis. A constitutively produced family of five triheme cytochromes (PpcA-E) from the bacterium Geobacter sulfurreducens plays a crucial role in extracellular electron transfer, a hallmark that permits this bacterium to be explored for several biotechnological applications. Two members of this family (PpcA and PpcD) couple electron/proton transfer in the physiological pH range, a feature not shared with PpcB and PpcE. That ability is crucial for G. sulfurreducens' growth in Fe(III)-reducing habitats since extra contributors to the electrochemical gradient are needed. It was postulated that the redox-Bohr effect is determined by the nature of residue 6, a leucine in PpcA/PpcD and a phenylalanine in PpcB/PpcE. To confirm this hypothesis, Phe6 was replaced by leucine in PpcB and PpcE. The functional properties of these mutants were investigated by NMR and UV-visible spectroscopy to assess their capability to couple electron/proton transfer in the physiological pH range. The results obtained showed that the mutants have an increased redox-Bohr effect and are now capable of coupling electron/proton transfer. This confirms the determinant role of the nature of residue 6 in the modulation of the redox-Bohr effect in this family of cytochromes, opening routes to engineer Geobacter cells with improved biomass production.


Assuntos
Proteínas de Bactérias/metabolismo , Citocromos/metabolismo , Geobacter/metabolismo , Heme/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Citocromos/química , Citocromos/genética , Transporte de Elétrons/genética , Elétrons , Geobacter/genética , Heme/química , Concentração de Íons de Hidrogênio , Modelos Moleculares , Estrutura Molecular , Mutação , Oxirredução , Conformação Proteica , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Prótons , Espectrofotometria/métodos , Termodinâmica
9.
Biomed Res Int ; 2021: 5574789, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34046497

RESUMO

Cytochrome (CYP) enzymes catalyze the metabolic reactions of endogenous and exogenous compounds. The superfamily of enzymes is found across many organisms, regardless of type, except for plants. Information was gathered about CYP2D enzymes through protein sequences of humans and other organisms. The secondary structure was predicted using the SOPMA. The structural and functional study of human CYP2D was conducted using ProtParam, SOPMA, Predotar 1.03, SignalP, TMHMM 2.0, and ExPASy. Most animals shared five central motifs according to motif analysis results. The tertiary structure of human CYP2D, as well as other animal species, was predicted by Phyre2. Human CYP2D proteins are heavily conserved across organisms, according to the findings. This indicates that they are descended from a single ancestor. They calculate the ratio of alpha-helices to extended strands to beta sheets to random coils. Most of the enzymes are alpha-helix, but small amounts of the random coil were also found. The data were obtained to provide us with a better understanding of mammalian proteins' functions and evolutionary relationships.


Assuntos
Citocromos/química , Citocromos/classificação , Filogenia , Proteínas/química , Sequência de Aminoácidos , Animais , Biologia Computacional/métodos , Simulação por Computador , Sistema Enzimático do Citocromo P-450/química , Sistema Enzimático do Citocromo P-450/classificação , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Citocromos/genética , Citocromos/metabolismo , Humanos , Ligantes , Camundongos , Modelos Moleculares , Conformação Proteica em alfa-Hélice , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Proteínas/classificação , Proteínas/genética , Alinhamento de Sequência , Software
10.
J Biol Chem ; 296: 100711, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33915126

RESUMO

Geobacter bacteria are able to transfer electrons to the exterior of the cell and reduce extracellular electron acceptors including toxic/radioactive metals and electrode surfaces, with potential applications in bioremediation or electricity harvesting. The triheme c-type cytochrome PpcA from Geobacter metallireducens plays a crucial role in bridging the electron transfer from the inner to the outer membrane, ensuring an effective extracellular electron transfer. This cytochrome shares 80% identity with PpcA from Geobacter sulfurreducens, but their redox properties are markedly different, thus determining the distinctive working redox potential ranges in the two bacteria. PpcA from G. metallireducens possesses two extra aromatic amino acids (Phe-6 and Trp-45) in its hydrophobic heme core, whereas PpcA from G. sulfurreducens has a leucine and a methionine in the equivalent positions. Given the different nature of these residues in the two cytochromes, we have hypothesized that the extra aromatic amino acids could be partially responsible for the observed functional differences. In this work, we have replaced Phe-6 and Trp-45 residues by their nonaromatic counterparts in PpcA from G. sulfurreducens. Using redox titrations followed by UV-visible and NMR spectroscopy we observed that residue Trp-45 shifted the redox potential range 33% toward that of PpcA from G. sulfurreducens, whereas Phe-6 produced a negligible effect. For the first time, it is shown that the inclusion of an aromatic residue at the heme core can modulate the working redox range in abundant periplasmic proteins, paving the way to engineer bacterial strains for optimal microbial bioelectrochemical applications.


Assuntos
Citocromos/química , Citocromos/metabolismo , Geobacter/citologia , Geobacter/enzimologia , Heme , Periplasma/enzimologia , Interações Hidrofóbicas e Hidrofílicas , Cinética , Modelos Moleculares , Oxirredução , Domínios Proteicos
11.
Biochemistry (Mosc) ; 86(1): 8-21, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-33705278

RESUMO

Bacillus subtilis serves as a model Gram-positive bacterium and an experimental system for research on respiratory enzymes. This review presents the heme proteins currently known for the well-characterized laboratory strain B. subtilis 168. It focuses on advances in research made during the last three decades concerning the function and composition of the cytochrome bc complex, terminal oxidases, and succinate:menaquinone oxidoreductase. The aerobic respiratory system of strain 168 seems representative for the species B. subtilis, as determined by the cytochrome composition of the undomesticated strain B. subtilis NCIB 3610 and a set of constructed cytochrome-deficient mutants of this strain. Unexplained and unsettled aspects of the molecular biology of respiratory cytochromes in B. subtilis are highlighted in the review.


Assuntos
Bacillus subtilis/metabolismo , Citocromos/metabolismo , Proteínas de Bactérias , Citocromos/genética , Transporte de Elétrons
12.
Appl Environ Microbiol ; 87(10)2021 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-33741623

RESUMO

Extracellular electron transfer (EET) is an important biological process in microbial physiology as found in dissimilatory metal oxidation/reduction and interspecies electron transfer in syntrophy in natural environments. EET also plays a critical role in microorganisms relevant to environmental biotechnology in metal-contaminated areas, metal corrosion, bioelectrochemical systems, and anaerobic digesters. Geobacter species exist in a diversity of natural and artificial environments. One of the outstanding features of Geobacter species is the capability of direct EET with solid electron donors and acceptors, including metals, electrodes, and other cells. Therefore, Geobacter species are pivotal in environmental biogeochemical cycles and biotechnology applications. Geobacter sulfurreducens, a representative Geobacter species, has been studied for direct EET as a model microorganism. G. sulfurreducens employs electrically conductive pili (e-pili) and c-type cytochromes for the direct EET. The biological function and electronics applications of the e-pili have been reviewed recently, and this review focuses on the cytochromes. Geobacter species have an unusually large number of cytochromes encoded in their genomes. Unlike most other microorganisms, Geobacter species localize multiple cytochromes in each subcellular fraction, outer membrane, periplasm, and inner membrane, as well as in the extracellular space, and differentially utilize these cytochromes for EET with various electron donors and acceptors. Some of the cytochromes are functionally redundant. Thus, the EET in Geobacter is complicated. Geobacter coordinates the cytochromes with other cellular components in the elaborate EET system to flourish in the environment.


Assuntos
Citocromos/metabolismo , Geobacter/metabolismo , Membrana Externa Bacteriana/metabolismo , Transporte de Elétrons , Membranas Intracelulares/metabolismo , Periplasma/metabolismo
13.
Nat Commun ; 12(1): 1104, 2021 02 17.
Artigo em Inglês | MEDLINE | ID: mdl-33597527

RESUMO

Photosynthetic electron transfers occur through multiple components ranging from small soluble proteins to large integral membrane protein complexes. Co-crystallization of a bacterial photosynthetic electron transfer complex that employs weak hydrophobic interactions was achieved by using high-molar-ratio mixtures of a soluble donor protein (high-potential iron-sulfur protein, HiPIP) with a membrane-embedded acceptor protein (reaction center, RC) at acidic pH. The structure of the co-complex offers a snapshot of a transient bioenergetic event and revealed a molecular basis for thermodynamically unfavorable interprotein electron tunneling. HiPIP binds to the surface of the tetraheme cytochrome subunit in the light-harvesting (LH1) complex-associated RC in close proximity to the low-potential heme-1 group. The binding interface between the two proteins is primarily formed by uncharged residues and is characterized by hydrophobic features. This co-crystal structure provides a model for the detailed study of long-range trans-protein electron tunneling pathways in biological systems.


Assuntos
Proteínas de Bactérias/química , Chromatiaceae/metabolismo , Proteínas Ferro-Enxofre/química , Complexos de Proteínas Captadores de Luz/química , Fotossíntese , Complexo de Proteínas do Centro de Reação Fotossintética/química , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Cristalização , Citocromos/química , Citocromos/metabolismo , Transporte de Elétrons , Heme/análogos & derivados , Heme/química , Heme/metabolismo , Proteínas Ferro-Enxofre/metabolismo , Complexos de Proteínas Captadores de Luz/metabolismo , Modelos Moleculares , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Conformação Proteica , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
14.
FEBS J ; 288(2): 382-404, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32530125

RESUMO

The Apicomplexa phylum groups important human and animal pathogens that cause severe diseases, encompassing malaria, toxoplasmosis, and cryptosporidiosis. In common with most organisms, apicomplexans rely on heme as cofactor for several enzymes, including cytochromes of the electron transport chain. This heme derives from de novo synthesis and/or the development of uptake mechanisms to scavenge heme from their host. Recent studies have revealed that heme synthesis is essential for Toxoplasma gondii tachyzoites, as well as for the mosquito and liver stages of Plasmodium spp. In contrast, the erythrocytic stages of the malaria parasites rely on scavenging heme from the host red blood cell. The unusual heme synthesis pathway in Apicomplexa spans three cellular compartments and comprises enzymes of distinct ancestral origin, providing promising drug targets. Remarkably given the requirement for heme, T. gondii can tolerate the loss of several heme synthesis enzymes at a high fitness cost, while the ferrochelatase is essential for survival. These findings indicate that T. gondii is capable of salvaging heme precursors from its host. Furthermore, heme is implicated in the activation of the key antimalarial drug artemisinin. Recent findings established that a reduction in heme availability corresponds to decreased sensitivity to artemisinin in T. gondii and Plasmodium falciparum, providing insights into the possible development of combination therapies to tackle apicomplexan parasites. This review describes the microeconomics of heme in Apicomplexa, from supply, either from de novo synthesis or scavenging, to demand by metabolic pathways, including the electron transport chain.


Assuntos
Cryptosporidium/metabolismo , Citocromos/metabolismo , Heme/metabolismo , Plasmodium berghei/metabolismo , Plasmodium falciparum/metabolismo , Proteínas de Protozoários/metabolismo , Toxoplasma/metabolismo , Animais , Anti-Infecciosos/farmacologia , Artemisininas/farmacologia , Cryptosporidium/efeitos dos fármacos , Cryptosporidium/genética , Cryptosporidium/crescimento & desenvolvimento , Citocromos/química , Citocromos/genética , Eritrócitos/metabolismo , Eritrócitos/parasitologia , Ferroquelatase/genética , Ferroquelatase/metabolismo , Expressão Gênica , Heme/química , Heme/genética , Interações Hospedeiro-Patógeno/genética , Humanos , Estágios do Ciclo de Vida/genética , Redes e Vias Metabólicas/genética , Plasmodium berghei/efeitos dos fármacos , Plasmodium berghei/genética , Plasmodium berghei/crescimento & desenvolvimento , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/genética , Plasmodium falciparum/crescimento & desenvolvimento , Proteínas de Protozoários/genética , Toxoplasma/efeitos dos fármacos , Toxoplasma/genética , Toxoplasma/crescimento & desenvolvimento
15.
Cell Biochem Biophys ; 79(1): 3-10, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32989571

RESUMO

Quinones are found in the lipid membranes of prokaryotes like E. coli and cyanobacteria, and are also abundant in eukaryotic mitochondria and chloroplasts. They are intricately involved in the reaction mechanism of redox phosphorylations. In the Mitchellian chemiosmotic school of thought, membrane-lodged quinones are perceived as highly mobile conveyors of two-electron equivalents from the first leg of Electron Transport Chain (ETC) to the 'second pit-stop' of Cytochrome bc1 or b6f complex (CBC), where they undergo a regenerative 'Q-cycle'. In Manoj's murburn mechanism, the membrane-lodged quinones are perceived as relatively slow-moving one- or two- electron donors/acceptors, enabling charge separation and the CBC resets a one-electron paradigm via 'turbo logic'. Herein, we compare various purviews of the two mechanistic schools with respect to: constraints in mobility, protons' availability, binding of quinones with proteins, structural features of the protein complexes, energetics of reaction, overall reaction logic, etc. From various perspectives, the murburn mechanism appeals as a viable alternative explanation well-rooted in thermodynamics/kinetics and one which lends adequate structure-function correlations for the roles of quinones, lipid membrane and associated proteins.


Assuntos
Cloroplastos/metabolismo , Lipídeos de Membrana/metabolismo , Mitocôndrias/metabolismo , Quinonas/metabolismo , Trifosfato de Adenosina/metabolismo , Sítios de Ligação , Citocromos/metabolismo , Transporte de Elétrons , Metabolismo Energético , Escherichia coli/metabolismo , Cinética , Bicamadas Lipídicas/química , Osmose , Oxirredução , Fosforilação Oxidativa , Oxigênio/química , Fotossíntese , Prótons , Quinonas/química , Relação Estrutura-Atividade , Termodinâmica
16.
Water Res ; 190: 116721, 2021 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-33326896

RESUMO

Intermittent (every other day) microaerobic [picomolar oxygen by oxidation-reduction potential (ORP) set at +25 mV above anaerobic baseline] digestion of lignocellulosic biomass showed higher digestibility and better stability at a high organic loading rate (OLR) of 5 g volatile solids (VS)/L/d than that under strict anaerobic conditions. However, the microbial mechanisms supporting the delicate balance under microaeration remain underexplored. On the basis of our previous findings that microbial communities in replicate experiments were dominated by strains of the genus Proteiniphilum but contained diverse taxa of methanogenic archaea, here we recovered related genomes and reconstructed the putative metabolic pathways using a genome-centric metagenomic approach. The highly enriched Proteiniphilum strains were identified as efficient cellulolytic facultative bacterium, which directly degraded lignocellulose to carbon dioxide, formate, and acetate via aerobic respiration and anaerobic fermentation, alternatively. Moreover, high oxygen affinity cytochromes, bd-type terminal oxidases, in Proteiniphilum strains were found to be closely associated with such picomolar oxygen conditions, which has long been overlooked in anaerobic digestion. Furthermore, hydrogenotrophic methanogenesis was the prevalent pathway for methane production while Methanosarcina, Methanobrevibacter, and Methanocorpusculum were the dominant methanogens in the replicate experiments. Importantly, the two functional groups, namely cellulolytic facultative Proteiniphilum strains and methanogens, encoded various antioxidant enzymes. Energy-dependent reactive oxygen species (ROS) scavengers (superoxide reductase (SOR) and rubrerythrin (rbr) were ubiquitously present in different methanogenic taxa in response to replicate-specific ORP levels (-470, -450 and -475 mV). Collectively, cytochrome bd oxidase and ROS defenders may play roles in improving the digestibility and stability observed in intermittent microaerobic digestion.


Assuntos
Citocromos , Metano , Anaerobiose , Biomassa , Reatores Biológicos , Citocromos/metabolismo , Lignina , Espécies Reativas de Oxigênio
17.
New Phytol ; 229(3): 1810-1821, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-32984969

RESUMO

Temperature dependence of plant respiratory O2 -consumption has been empirically described by the Arrhenius equation. The slope of the Arrhenius plot (which is proportional to activation energy) sometimes deviates from a constant value. We conducted kinetic model simulations of mitochondrial electron flow dynamics to clarify factors affecting the shape of the Arrhenius plot. We constructed a kinetic model of respiration in which competitive O2 -consumption by the cytochrome pathway (CP) and the alternative pathway (AP) were considered, and we used this model to describe the temperature dependence of respiratory O2 -consumption of Arabidopsis. The model indicated that the electron partitioning and activation energy differences between CP and AP were reflected in the slope and magnitude of the dependent variables of the Arrhenius plot. When the electron partitioning and activation energies of CP and AP were constant with temperature change, our model suggested that the Arrhenius plot would be almost linear. When the electron partitioning or activation energy of CP, or both, rapidly changed with temperature, the Arrhenius plot deviated from linearity, as reported in previous experimental studies. Our simulation analysis quantitatively linked the kinetic model parameters with physiological mechanisms underlying the instantaneous temperature dependence of plant respiration rate.


Assuntos
Arabidopsis , Citocromos , Arabidopsis/metabolismo , Citocromos/metabolismo , Transporte de Elétrons , Cinética , Temperatura
18.
Curr Opin Chem Biol ; 59: 193-201, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33070100

RESUMO

Extracellular electron transfer via filamentous protein appendages called 'microbial nanowires' has long been studied in Geobacter and other bacteria because of their crucial role in globally-important environmental processes and their applications for bioenergy, biofuels, and bioelectronics. Thousands of papers thought these nanowires as pili without direct evidence. Here, we summarize recent discoveries that could help resolve two decades of confounding observations. Using cryo-electron microscopy with multimodal functional imaging and a suite of electrical, biochemical, and physiological studies, we find that rather than pili, nanowires are composed of cytochromes OmcS and OmcZ that transport electrons via seamless stacking of hemes over micrometers. We discuss the physiological need for two different nanowires and their potential applications for sensing, synthesis, and energy production.


Assuntos
Proteínas de Bactérias/metabolismo , Citocromos/metabolismo , Fímbrias Bacterianas/metabolismo , Bactérias Gram-Negativas/metabolismo , Proteínas de Bactérias/ultraestrutura , Citocromos/ultraestrutura , Transporte de Elétrons , Fímbrias Bacterianas/ultraestrutura , Geobacter/metabolismo , Geobacter/ultraestrutura , Bactérias Gram-Negativas/ultraestrutura , Modelos Moleculares , Nanofios/ultraestrutura
19.
Biochemistry (Mosc) ; 85(8): 955-965, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-33045956

RESUMO

The concept of "electric cables" involved in bioenergetic processes of a living cell was proposed half a century ago [Skulachev, V. P. (1971) Curr. Top. Bioenerg., Elsevier, pp. 127-190]. For many decades, only cell membrane structures have been considered as probable pathways for the electric current, namely, for the transfer of transmembrane electrochemical potential. However, the last ten to fifteen years have brought the discovery of bacterial "electric cables" of a new type. In 2005, "nanowires" conducting electric current over distances of tens of micrometers were discovered in metal- and sulphate-reducing bacteria [Reguera, G. et al. (2005) Nature, 435, pp. 1098-1101]. The next five years have witnessed the discovery of microbial electric currents over centimeter distances [Nielsen, L. P. et al. (2010) Nature, 463, 1071-1074]. This new group of bacteria allowing electric currents to flow over macroscopic distances was later called cable bacteria. Nanowires and conductive structures of cable bacteria serve to solve a special problem of membrane bioenergetics: they connect two redox half-reactions. In other words, unlike membrane "cables", their function is electron transfer in the course of oxidative phosphorylation for the generation of membrane energy rather than of the end-product. The most surprising is the protein nature of these cables (at least of some of them) indicated by recent data, since no protein wires for the long-distance electron transport had been previously known in living systems.


Assuntos
Condutividade Elétrica , Geobacter/citologia , Geobacter/metabolismo , Citocromos/metabolismo , Transporte de Elétrons , Elétrons , Proteínas de Fímbrias/metabolismo , Fímbrias Bacterianas/química , Fímbrias Bacterianas/metabolismo , Potenciais da Membrana/fisiologia , Oxirredução , Fosforilação Oxidativa , Conformação Proteica
20.
Appl Environ Microbiol ; 86(22)2020 10 28.
Artigo em Inglês | MEDLINE | ID: mdl-32917753

RESUMO

Proteins that oxidize extracellular substrates in Gram-positive bacteria are poorly understood. Ferrimicrobium acidiphilum is an actinobacterium that respires aerobically on extracellular ferrous ions at pH 1.5. In situ absorbance measurements were conducted on turbid suspensions of intact Fm. acidiphilum using an integrating cavity absorption meter designed for that purpose. Initial velocity kinetic studies monitored the appearance of product ferric ions in the presence of catalytic quantities of cells. Cell-catalyzed iron oxidation obeyed the Michaelis-Menten equation with Km and V max values of 71 µM and 0.29 fmol/min/cell, respectively. Limited-turnover kinetic studies were conducted with higher concentrations of cells to detect and monitor changes in the absorbance properties of cellular redox proteins when the cells were exposed to limited quantities of soluble reduced iron. A single a-type cytochrome with reduced absorbance peaks at 448 and 605 nm was the only redox-active chromophore that was visible as the cells respired aerobically on iron. The reduced cytochrome 605 exhibited mathematical and correlational properties that were consistent with the hypothesis that oxidation of the cytochrome constituted the rate-limiting step in the aerobic respiratory process, with a turnover number of 35 ± 2 s-1 Genomic and proteomic analyses showed that Fm. acidiphilum could and did express only two a-type heme copper terminal oxidases. Cytochrome 605 was associated with the terminal oxidase gene that is located between nucleotides 31,090 and 33,039, inclusive, in the annotated circular genome of this bacterium.IMPORTANCE The identities and functions of proteins involved in aerobic respiration on extracellular ferrous ions at acidic pH are poorly understood in the four phyla of Gram-positive eukaryotes and archaea where such activities occur. In situ absorbance measurements were conducted on Fm. acidiphilum as it respired on extracellular iron using an integrating cavity absorption meter that permitted accurate optical measurements in turbid suspensions of the intact bacterium under physiological conditions. The significance of these measurements is that they permitted a direct spectrophotometric examination of the extents and rates of biological electron transfer events in situ under noninvasive physiological conditions without disrupting the complexity of the live cellular environment. One thing is certain: one way to understand how a protein functions in an intact organism is to actually observe that protein as it functions in the intact organism. This paper provides an example of just such an observation.


Assuntos
Actinobacteria/metabolismo , Proteínas de Bactérias/metabolismo , Citocromos/metabolismo , Ferro/metabolismo , Aerobiose , Oxirredução
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