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1.
J Biosci Bioeng ; 132(6): 569-574, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34518108

RESUMO

Gas fermentation is a promising biological process for the conversion of CO2 or syngas into valuable chemicals. Homoacetogens are microorganisms growing autotrophically using CO2 and H2 or CO and metabolizing them to form acetate coupled with energy conservation. The challenge in the metabolic engineering of the homoacetogens is divergence of the acetate formation, whose intermediate is acetyl-CoA, to a targeted chemical with sufficient production of adenosine triphosphate (ATP). In this study, we report that an engineered strain of the thermophilic homoacetogen Moorella thermoacetica, in which a pool of acetyl-CoA is diverted to ethanol without ATP production, can maintain autotrophic growth on syngas. We estimated the ATP production in the engineered strains under different gaseous compositions by considering redox-balanced metabolism for ethanol and acetate formation. The culture test showed that the combination of retaining a level of acetate production and supplying the energy-rich CO allowed maintenance of the autotrophic growth during ethanol production. In contrast, autotrophy was collapsed by complete elimination of the acetate pathway or supplementation of H2-CO2. We showed that the intracellular level of ATP was significantly lowered on H2-CO2 in consistent with the incompetence. In the meantime, the complete disruption of the acetate pathway resulted in the redox imbalance to produce ethanol from CO, albeit a small loss in the ATP production. Thus, preservation of a fraction of acetate formation is required to maintain sufficient ATP and balanced redox in CO-containing gases for ethanol production.


Assuntos
Etanol , Moorella , Acetatos , Processos Autotróficos , Moorella/genética
2.
Water Res ; 197: 117081, 2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-33813170

RESUMO

Engineered nanoparticles are observed to be released into the environment and ended up in wastewater treatment plants. It has been reported that these nanoparticles in sewage might have a toxic effect on microorganisms, and thus affect anaerobic microbial fermentation. However, the mechanisms involved in nanoparticles-induced effects on the anaerobic acidification process and its related bacterial metabolism are still unclear. This work indicated that copper nanoparticles (Cu NPs) were able to cause cell membrane oxidative damage and inhibit the growth and metabolism of Moorella thermoacetica (a model acetogen). The OD600 and acetic acid production of M. thermoacetica in the presence of 1 mg/L of Cu NPs were decreased to 29.2% and 40.7% of the control, respectively. The key mechanism of the inhibitory effect was governed by the fact that Cu NPs significantly reduced the glucose consumption, and led to the decreased pyruvate metabolism levels. Additionally, Cu NPs inhibited the gene expressions and catalytic activities of the key enzymes related to acetic acid production. It was identified that the relative activities of phosphofructokinase, pyruvate kinase, phosphotransacetylase, and acetate kinase of M. thermoacetica in the presence of 1 mg/L of Cu NPs decreased to only 70.1%, 69.3%, 50.1%, and 65.2% of the control, respectively. These results demonstrated that the release of Cu NPs in the environment could pose risks to anaerobic fermentation processes via regulating microbial transcriptional response and enzyme activity.


Assuntos
Nanopartículas Metálicas , Nanopartículas , Anaerobiose , Cobre , Fermentação , Moorella
3.
Curr Microbiol ; 78(5): 1903-1913, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33786643

RESUMO

In this study, using a metagenomic approach, we explore the bacterial diversity of compost sites categorized based on their ambient temperatures. The two sites were Reckong Peo in the lower Himalayas and Tambaram in the southern region of the country, namely, CPR and CT. Following assembly of the raw reads from shotgun metagenomics, similarity hits were generated using NCBI BLAST + and SILVA database. A total of 1463 and 1483 species were annotated from CPR and CT. A species-level annotation was performed using a python-based literature search pipeline revealing their growth characteristics. Thermophiles Thermomonospora curvata and Thermus scotoductus were among the prominent species in CT. CPR too was seen abundant with Acidothermus cellulolyticus and Moorella thermoacetica, constituting 10% of the population. Nearly 3% of the identified species in the site CPR were psychrophilic. Although found higher in CPR, psychrophilic species were identified in CT too. Flavobacterium and Psychrobacter spp. were present in both sites without any significant changes in their relative distribution contrary to the thermophilic species abundance (z = - 4.3). Akin to the sequenced samples, database-derived metagenomes also showed similar distribution of thermophiles and psychrophiles. Identifying such peculiar prevalence of extremophiles can be central to understanding extended growth temperatures.


Assuntos
Compostagem , Metagenômica , Actinobacteria , Moorella , Temperatura , Thermus
4.
Sci Rep ; 11(1): 2139, 2021 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-33495538

RESUMO

Biohybrids composed of microorganisms and nanoparticles have emerged as potential systems for bioenergy and high-value compound production from CO2 and light energy, yet the cellular and metabolic processes within the biological component of this system are still elusive. Here we dissect the biohybrid composed of the anaerobic acetogenic bacterium Moorella thermoacetica and cadmium sulphide nanoparticles (CdS) in terms of physiology, metabolism, enzymatics and transcriptomic profiling. Our analyses show that while the organism does not grow on L-cysteine, it is metabolized to acetate in the biohybrid system and this metabolism is independent of CdS or light. CdS cells have higher metabolic activity, despite an inhibitory effect of Cd2+ on key enzymes, because of an intracellular storage compound linked to arginine metabolism. We identify different routes how cysteine and its oxidized form can be innately metabolized by the model acetogen and what intracellular mechanisms are triggered by cysteine, cadmium or blue light.


Assuntos
Carbono/metabolismo , Cisteína/metabolismo , Metabolismo Energético , Acetatos/metabolismo , Transporte Biológico/efeitos dos fármacos , Cádmio/farmacologia , Isótopos de Carbono , Misturas Complexas , Cisteína/farmacologia , Metabolismo Energético/efeitos dos fármacos , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Luz , Espectroscopia de Ressonância Magnética , Moorella/genética , Moorella/crescimento & desenvolvimento , Moorella/efeitos da radiação , Moorella/ultraestrutura , Oxirredução , Transcriptoma/genética
5.
Biochim Biophys Acta Bioenerg ; 1862(1): 148330, 2021 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-33080205

RESUMO

Clostridium autoethanogenum, the bacterial model for biological conversion of waste gases into biofuels, grows under extreme carbon-monoxide (CO) concentrations. The strictly anaerobic bacterium derives its entire cellular energy and carbon from this poisonous gas, therefore requiring efficient molecular machineries for CO-conversion. Here, we structurally and biochemically characterized the key enzyme of the CO-converting metabolism: the CO-dehydrogenase/Acetyl-CoA synthase (CODH/ACS). We obtained crystal structures of natively isolated complexes from fructose-grown and CO-grown C. autoethanogenum cultures. Both contain the same isoforms and if the overall structure adopts the classic α2ß2 architecture, comparable to the model enzyme from Moorella thermoacetica, the ACS binds a different position on the CODH core. The structural characterization of a proteolyzed complex and the conservation of the binding interface in close homologs rejected the possibility of a crystallization artefact. Therefore, the internal CO-channeling system, critical to transfer CO generated at the C-cluster to the ACS active site, drastically differs in the complex from C. autoethanogenum. The 1.9-Å structure of the CODH alone provides an accurate picture of the new CO-routes, leading to the ACS core and reaching the surface. Increased gas accessibility would allow the simultaneous CO-oxidation and acetyl-CoA production. Biochemical experiments showed higher flexibility of the ACS subunit from C. autoethanogenum compared to M. thermoacetica, albeit monitoring similar CO-oxidation and formation rates. These results show a reshuffling of internal CO-tunnels during evolution of these Firmicutes, putatively leading to a bidirectional complex that ensure a high flux of CO-conversion toward energy conservation, acting as the main cellular powerplant.


Assuntos
Acetilcoenzima A/química , Aldeído Oxirredutases/química , Proteínas de Bactérias/química , Monóxido de Carbono/química , Clostridium/enzimologia , Complexos Multienzimáticos/química , Acetilcoenzima A/metabolismo , Aldeído Oxirredutases/metabolismo , Proteínas de Bactérias/metabolismo , Monóxido de Carbono/metabolismo , Cristalografia por Raios X , Moorella/enzimologia , Complexos Multienzimáticos/metabolismo , Oxirredução , Estrutura Quaternária de Proteína
6.
Structure ; 29(1): 43-49.e3, 2021 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-32937101

RESUMO

The Ni-Fe-S-containing A-cluster of acetyl-coenzyme A (CoA) synthase (ACS) assembles acetyl-CoA from carbon monoxide (CO), a methyl group (CH3+), and CoA. To accomplish this feat, ACS must bind CoA and interact with two other proteins that contribute the CO and CH3+, respectively: CO dehydrogenase (CODH) and corrinoid Fe-S protein (CFeSP). Previous structural data show that, in the model acetogen Moorella thermoacetica, domain 1 of ACS binds to CODH such that a 70-Å-long internal channel is created that allows CO to travel from CODH to the A-cluster. The A-cluster is largely buried and is inaccessible to CFeSP for methylation. Here we use electron microscopy to capture multiple snapshots of ACS that reveal previously uncharacterized domain motion, forming extended and hyperextended structural states. In these structural states, the A-cluster is accessible for methylation by CFeSP.


Assuntos
Aldeído Oxirredutases/química , Proteínas de Bactérias/química , Complexos Multienzimáticos/química , Aldeído Oxirredutases/metabolismo , Proteínas de Bactérias/metabolismo , Ferro/química , Ferro/metabolismo , Simulação de Dinâmica Molecular , Moorella/enzimologia , Complexos Multienzimáticos/metabolismo , Níquel/química , Níquel/metabolismo , Domínios Proteicos , Enxofre/química , Enxofre/metabolismo
7.
Environ Sci Pollut Res Int ; 28(10): 11904-11914, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32048194

RESUMO

Valorisation of organic wastes to produce industrially relevant commodity products is a sustainable, cost-effective and viable alternative providing a green platform for chemical production while simultaneously leading to waste disposal management. In the present study, organic wastes such as agricultural residue-derived sugars, oilseed meals, poultry waste and molasses were used for substituting expensive organic fermentation medium components. Moorella thermoacetica and Aurantiochytrium limacinum were adapted on these waste-derived hydrolysates to produce high volume-low value products such as bio-acetic acid (80% theoretical yields) and oil-rich fish/animal feed (more than 85% dry cell weight as compared with conventional nutrient sources) respectively. Use of these waste-derived nutrients led to ~ 75% and ~ 90% reduction in media cost for acetic acid and oil-rich biomass production respectively as compared with that of traditionally used high-priced medium components. The strategy will assist in the cost reduction for high volume-low value products while also ensuring waste recovery.


Assuntos
Moorella , Estramenópilas , Animais , Biomassa , Fermentação , Resíduos
8.
Inorg Chem ; 59(20): 15167-15179, 2020 Oct 19.
Artigo em Inglês | MEDLINE | ID: mdl-33017144

RESUMO

The biological synthesis of acetyl-coenzyme A (acetyl-CoA), catalyzed by acetyl-CoA synthase (ACS), is of biological significance and chemical interest acting as a source of energy and carbon. The catalyst contains an unusual hexa-metal cluster with two nickel ions and a [Fe4S4] cluster. DFT calculations have been performed to investigate the ACS reaction mechanism starting from three different oxidation states (+2, +1, and 0) of Nip, the nickel proximal to [Fe4S4]. The results indicate that the ACS reaction proceeds first through a methyl radical transfer from cobalamin (Cbl) to Nip randomly accompanying with the CO binding. After that, C-C bond formation occurs between the Nip-bound methyl and CO, forming Nip-acetyl. The substrate CoA-S- then binds to Nip, allowing C-S bond formation between the Nip-bound acetyl and CoA-S-. Methyl transfer is rate-limiting with a barrier of ∼14 kcal/mol, which does not depend on the presence or absence of CO. Both the Nip2+ and Nip1+ states are chemically capable of catalyzing the ACS reaction independent of the state (+2 or +1) of the [Fe4S4] cluster. The [Fe4S4] cluster is not found to affect the steps of methyl transfer and C-C bond formation but may be involved in the C-S bond formation depending on the detailed mechanism chosen. An ACS active site containing a Nip(0) state could not be obtained. Optimizations always led to a Nip1+ state coupled with [Fe4S4]1+. The calculations show a comparable activity for Nip1+/[Fe4S4]1+, Nip1+/[Fe4S4]2+, and Nip2+/[Fe4S4]2+. The results here give significant insights into the chemistry of the important ACS reaction.


Assuntos
Acetato-CoA Ligase/química , Proteínas de Bactérias/química , Catálise , Teoria da Densidade Funcional , Firmicutes/enzimologia , Proteínas Ferro-Enxofre/química , Modelos Químicos , Moorella/enzimologia , Níquel/química , Oxirredução , Vitamina B 12/análogos & derivados , Vitamina B 12/química
9.
J Biol Chem ; 295(31): 10522-10534, 2020 07 31.
Artigo em Inglês | MEDLINE | ID: mdl-32503839

RESUMO

Vitamin B12 and other cobamides are essential cofactors required by many organisms and are synthesized by a subset of prokaryotes via distinct aerobic and anaerobic routes. The anaerobic biosynthesis of 5,6-dimethylbenzimidazole (DMB), the lower ligand of vitamin B12, involves five reactions catalyzed by the bza operon gene products, namely the hydroxybenzimidazole synthase BzaAB/BzaF, phosphoribosyltransferase CobT, and three methyltransferases, BzaC, BzaD, and BzaE, that conduct three distinct methylation steps. Of these, the methyltransferases that contribute to benzimidazole lower ligand diversity in cobamides remain to be characterized, and the precise role of the bza operon protein CobT is unclear. In this study, we used the bza operon from the anaerobic bacterium Moorella thermoacetica (comprising bzaA-bzaB-cobT-bzaC) to examine the role of CobT and investigate the activity of the first methyltransferase, BzaC. We studied the phosphoribosylation catalyzed by MtCobT and found that it regiospecifically activates 5-hydroxybenzimidazole (5-OHBza) to form the 5-OHBza-ribotide (5-OHBza-RP) isomer as the sole product. Next, we characterized the domains of MtBzaC and reconstituted its methyltransferase activity with the predicted substrate 5-OHBza and with two alternative substrates, the MtCobT product 5-OHBza-RP and its riboside derivative 5-OHBza-R. Unexpectedly, we found that 5-OHBza-R is the most favored MtBzaC substrate. Our results collectively explain the long-standing observation that the attachment of the lower ligand in anaerobic cobamide biosynthesis is regiospecific. In conclusion, we validate MtBzaC as a SAM:hydroxybenzimidazole-riboside methyltransferase (HBIR-OMT). Finally, we propose a new pathway for the synthesis and activation of the benzimidazolyl lower ligand in anaerobic cobamide biosynthesis.


Assuntos
Proteínas de Bactérias/metabolismo , Benzimidazóis/metabolismo , Cobamidas/biossíntese , Metiltransferases/metabolismo , Moorella/metabolismo , Pentosiltransferases/metabolismo , Anaerobiose , Proteínas de Bactérias/genética , Cobamidas/genética , Metilação , Metiltransferases/genética , Moorella/genética , Pentosiltransferases/genética
10.
Angew Chem Int Ed Engl ; 59(18): 7224-7229, 2020 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-32065712

RESUMO

An organic semiconductor-bacteria biohybrid photosynthetic system is used to efficiently realize CO2 reduction to produce acetic acid with the non-photosynthetic bacteria Moorella thermoacetica. Perylene diimide derivative (PDI) and poly(fluorene-co-phenylene) (PFP) were coated on the bacteria surface as photosensitizers to form a p-n heterojunction (PFP/PDI) layer, affording higher hole/electron separation efficiency. The π-conjugated semiconductors possess excellent light-harvesting ability and biocompatibility, and the cationic side chains of organic semiconductors could intercalate into cell membranes, ensuring efficient electron transfer to bacteria. Moorella thermoacetica can thus harvest photoexcited electrons from the PFP/PDI heterojunction, driving the Wood-Ljungdahl pathway to synthesize acetic acid from CO2 under illumination. The efficiency of this organic biohybrid is about 1.6 %, which is comparable to those of reported inorganic biohybrid systems.


Assuntos
Ácido Acético/metabolismo , Dióxido de Carbono/metabolismo , Moorella/metabolismo , Fármacos Fotossensibilizantes/metabolismo , Energia Solar , Ácido Acético/química , Dióxido de Carbono/química , Transporte de Elétrons , Fluorenos/química , Fluorenos/metabolismo , Imidas/química , Imidas/metabolismo , Estrutura Molecular , Moorella/citologia , Oxirredução , Perileno/análogos & derivados , Perileno/química , Perileno/metabolismo , Fármacos Fotossensibilizantes/química , Polímeros/química , Polímeros/metabolismo , Semicondutores , Propriedades de Superfície
11.
J Biosci Bioeng ; 129(2): 160-164, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31506242

RESUMO

Bioconversion from inexpensive renewable resource, such as biomass, to liquid fuel is one of the promising technologies to reduce the use of petroleum. We previously reported the genetically engineered Moorella thermoacetica could produce ethanol from the lignocellulosic feedstock. However, it was still unclear which carbon source in the substrate was preferentially consumed to produce ethanol. To identify the hierarchy of the sugar utilization during ethanol fermentation of this strain, we analyzed the sugar composition of lignocellulosic feedstock, and consumption rate of sugars during the fermentation process. The hydrolysates after acid pretreatment and enzymatic saccharification contained glucose, xylose, galactose, arabinose, and mannose. Time course data suggested that xylose was the most preferred carbon source among those sugars during ethanol fermentation. Ethanol yield was 0.40 ± 0.06 and 0.40 ± 0.12 g/g-total sugar, from lignocellulosic hydrolysates of Japanese cedar (Cryptomeria japonica) and rice straw (Oryza sativa), respectively. The results demonstrated that the genetically engineered M. thermoacetica is a promising candidate for thermophilic ethanol fermentation of lignocellulosic feedstocks, especially hemicellulosic sugars.


Assuntos
Etanol/metabolismo , Lignina/metabolismo , Moorella/metabolismo , Açúcares/metabolismo , Fermentação , Engenharia Genética , Temperatura Alta , Hidrólise , Moorella/genética
12.
Nat Commun ; 10(1): 3311, 2019 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-31427571

RESUMO

Genome-wide analysis of DNA methylation patterns using single molecule real-time DNA sequencing has boosted the number of publicly available methylomes. However, there is a lack of tools coupling methylation patterns and the corresponding methyltransferase genes. Here we demonstrate a high-throughput method for coupling methyltransferases with their respective motifs, using automated cloning and analysing the methyltransferases in vectors carrying a strain-specific cassette containing all potential target sites. To validate the method, we analyse the genomes of the thermophile Moorella thermoacetica and the mesophile Acetobacterium woodii, two acetogenic bacteria having substantially modified genomes with 12 methylation motifs and a total of 23 methyltransferase genes. Using our method, we characterize the 23 methyltransferases, assign motifs to the respective enzymes and verify activity for 11 of the 12 motifs.


Assuntos
Acetobacterium/enzimologia , Proteínas de Bactérias/metabolismo , Ensaios de Triagem em Larga Escala/métodos , Metiltransferases/metabolismo , Moorella/enzimologia , Acetobacterium/genética , Motivos de Aminoácidos/genética , Proteínas de Bactérias/genética , Metilação de DNA , DNA Bacteriano/metabolismo , Epigênese Genética , Genoma Bacteriano , Moorella/genética , Análise de Sequência de DNA
13.
PLoS One ; 14(6): e0216979, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31181074

RESUMO

An ancient enzyme family responsible for the catabolism of the prebiotic chemical cyanuric acid (1,3,5-triazine-2,4,6-triol) was recently discovered and is undergoing proliferation in the modern world due to industrial synthesis and dissemination of 1,3,5-triazine compounds. Cyanuric acid has a highly stabilized ring system such that bacteria require a unique enzyme with a novel fold and subtle active site construction to open the ring. Each cyanuric acid hydrolase monomer consists of three isostructural domains that coordinate and activate the three-fold symmetric substrate cyanuric acid for ring opening. We have now solved a series of X-ray structures of an engineered, thermostable cyanuric acid ring-opening enzyme at 1.51 ~ 2.25 Å resolution, including various complexes with the substrate, a tight-binding inhibitor, or an analog of the reaction intermediate. These structures reveal asymmetric interactions between the enzyme and bound ligands, a metal ion binding coupled to conformational changes and substrate binding important for enzyme stability, and distinct roles of the isostructural domains of the enzyme. The multiple conformations of the enzyme observed across a series of structures and corroborating biochemical data suggest importance of the structural dynamics in facilitating the substrate entry and the ring-opening reaction, catalyzed by a conserved Ser-Lys dyad.


Assuntos
Biocatálise , Hidrolases/química , Hidrolases/metabolismo , Moorella/enzimologia , Triazinas/metabolismo , Domínio Catalítico , Cristalografia por Raios X , Modelos Moleculares , Multimerização Proteica , Estrutura Quaternária de Proteína
14.
Inorg Chem ; 58(12): 7931-7938, 2019 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-31141352

RESUMO

Nickel-containing carbon monoxide (CO) dehydrogenase is an enzyme that catalyzes the important reversible carbon dioxide reduction. Several high-resolution structures have been determined at various stages of the reduction, which can be used as good starting points for the present computational study. The cluster model is used in combination with a systematic application of the density functional theory as recently described. The results are in very good agreement with experimental evidence. There are a few important results. To explain why the X-ray structure for the reduced Cred1 state has an empty site on nickel, it is here suggested that the cluster has been over-reduced by X-rays and is therefore not the desired reduced state, which instead contains a bound CO on nickel. After an additional reduction, a hydride bound to nickel is suggested to play a role. In order to obtain energetics in agreement with experiments, it is concluded that one sulfide bridge in the Ni-Fe cluster should be protonated. The best test of the accuracy obtained is to compare the computed rate for reduction using -0.6 V with that for oxidation using -0.3 V, where good agreement was obtained. Obtaining a mechanism that is easily reversible is another demanding aspect of the modeling. Nickel oscillates between nickel(II) and nickel(I), while nickel(0) never comes in.


Assuntos
Aldeído Oxirredutases/química , Monóxido de Carbono/química , Proteínas Ferro-Enxofre/química , Complexos Multienzimáticos/química , Níquel/química , Domínio Catalítico , Cristalografia por Raios X , Teoria da Densidade Funcional , Desulfovibrio vulgaris/enzimologia , Methanosarcina barkeri/enzimologia , Modelos Químicos , Moorella/enzimologia , Oxirredução , Rhodospirillum rubrum/enzimologia , Termodinâmica
15.
Faraday Discuss ; 215(0): 54-65, 2019 07 04.
Artigo em Inglês | MEDLINE | ID: mdl-30994635

RESUMO

Solar-driven conversion of carbon dioxide to value-added carbon products is an ambitious objective of ongoing research efforts. However, high overpotential, low selectivity and poor CO2 mass transfer plague purely inorganic electrocatalysts. In this instance, we can consider a class of biological organisms that have evolved to achieve CO2 fixation. We can harness and combine the streamlined CO2 fixation pathways of these whole organisms with the exceptional ability of semiconducting nanomaterials to harvest solar energy. A novel nanomaterial-biological interface has been pioneered in which light-capturing cadmium sulfide nanoparticles reside within individual organisms essentially powering biological CO2 fixation by solar energy. In order to further develop the photosensitized organism platform, more biocompatible photosensitizers and cytoprotective strategies are required as well as elucidation of charge transfer mechanisms. Here, we discuss the ability of gold nanoclusters to photosensitize a model acetogen effectively and biocompatibly. Additionally, we present innovative materials including two-dimensional metal organic framework sheets and alginate hydrogels to shield photosensitized cells. Finally, we delve into original work using transient absorption spectroscopy to inform on charge transfer mechanisms.


Assuntos
Dióxido de Carbono/química , Ouro/química , Nanopartículas Metálicas/química , Estruturas Metalorgânicas/química , Fármacos Fotossensibilizantes/química , Energia Solar , Ouro/metabolismo , Estruturas Metalorgânicas/metabolismo , Moorella/química , Moorella/metabolismo , Fármacos Fotossensibilizantes/metabolismo , Fotossíntese , Semicondutores
16.
Nat Metab ; 1(6): 643-651, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-32694804

RESUMO

Advanced bioproduct synthesis via reductive metabolism requires coordinating carbons, ATP and reducing agents, which are generated with varying efficiencies depending on metabolic pathways. Substrate mixtures with direct access to multiple pathways may optimally satisfy these biosynthetic requirements. However, native regulation favouring preferential use precludes cells from co-metabolizing multiple substrates. Here we explore mixed substrate metabolism and tailor pathway usage to synergistically stimulate carbon reduction. By controlled cofeeding of superior ATP and NADPH generators as 'dopant' substrates to cells primarily using inferior substrates, we circumvent catabolite repression and drive synergy in two divergent organisms. Glucose doping in Moorella thermoacetica stimulates CO2 reduction (2.3 g gCDW-1 h-1) into acetate by augmenting ATP synthesis via pyruvate kinase. Gluconate doping in Yarrowia lipolytica accelerates acetate-driven lipogenesis (0.046 g gCDW-1 h-1) by obligatory NADPH synthesis through the pentose cycle. Together, synergistic cofeeding produces CO2-derived lipids with 38% energy yield and demonstrates the potential to convert CO2 into advanced bioproducts. This work advances the systems-level control of metabolic networks and CO2 use, the most pressing and difficult reduction challenge.


Assuntos
Moorella/metabolismo , Yarrowia/metabolismo , Trifosfato de Adenosina/metabolismo , Ciclo do Ácido Cítrico/fisiologia , Glucose/metabolismo , NADP/metabolismo , Oxirredução , Via de Pentose Fosfato/fisiologia
17.
Biotechnol Bioeng ; 116(2): 294-306, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30267586

RESUMO

Synthesis gas (syngas) fermentation via the Wood-Ljungdahl pathway is receiving growing attention as a possible platform for the fixation of CO 2 and renewable production of fuels and chemicals. However, the pathway operates near the thermodynamic limit of life, resulting in minimal adenosine triphosphate (ATP) production and long doubling times. This calls into question the feasibility of producing high-energy compounds at industrially relevant levels. In this study, we investigated the possibility of co-utilizing nitrate as an inexpensive additional electron acceptor to enhance ATP production during H 2 -dependent growth of Clostridium ljungdahlii, Moorella thermoacetica, and Acetobacterium woodii. In contrast to other acetogens tested, growth rate and final biomass titer were improved for C. ljungdahlii growing on a mixture of H 2 and CO 2 when supplemented with nitrate. Transcriptomic analysis, 13 CO 2 labeling, and an electron balance were used to understand how electron flux was partitioned between CO 2 and nitrate. We further show that, with nitrate supplementation, the ATP/adenosine diphosphate (ADP) ratio and acetyl-CoA pools were increased by fivefold and threefold, respectively, suggesting that this strategy could be useful for the production of ATP-intensive heterologous products from acetyl-CoA. Finally, we propose a pathway for enhanced ATP production from nitrate and use this as a basis to calculate theoretical yields for a variety of products. This study demonstrates a viable strategy for the decoupling of ATP production from carbon dioxide fixation, which will serve to significantly improve the CO 2 fixation rate and the production metrics of other chemicals from CO 2 and H 2 in this host.


Assuntos
Acetobacterium/metabolismo , Dióxido de Carbono/metabolismo , Clostridium/metabolismo , Hidrogênio/metabolismo , Moorella/metabolismo , Nitratos/metabolismo , Acetobacterium/crescimento & desenvolvimento , Trifosfato de Adenosina/biossíntese , Ciclo do Carbono , Clostridium/crescimento & desenvolvimento , Análise do Fluxo Metabólico , Moorella/crescimento & desenvolvimento
18.
Proc Natl Acad Sci U S A ; 115(42): 10582-10587, 2018 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-30275326

RESUMO

We report a strategy to uniformly wrap Morella thermoacetica bacteria with a metal-organic framework (MOF) monolayer of nanometer thickness for cytoprotection in artificial photosynthesis. The catalytic activity of the MOF enclosure toward decomposition of reactive oxygen species (ROS) reduces the death of strictly anaerobic bacteria by fivefold in the presence of 21% O2, and enables the cytoprotected bacteria to continuously produce acetate from CO2 fixation under oxidative stress. The high definition of the MOF-bacteria interface involving direct bonding between phosphate units on the cell surface and zirconium clusters on MOF monolayer, provides for enhancement of life throughout reproduction. The dynamic nature of the MOF wrapping allows for cell elongation and separation, including spontaneous covering of the newly grown cell surface. The open-metal sites on the zirconium clusters lead to 600 times more efficient ROS decomposition compared with zirconia nanoparticles.


Assuntos
Citoproteção , Estruturas Metalorgânicas/química , Moorella/crescimento & desenvolvimento , Zircônio/química , Sobrevivência Celular , Estresse Oxidativo , Fotossíntese , Propriedades de Superfície
19.
Nat Plants ; 4(5): 241, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29725093
20.
Food Microbiol ; 73: 334-341, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29526221

RESUMO

Temperatures encountered in cannery allow growth of thermophilic spore-forming bacteria, including the strictly anaerobe Moorella thermoacetica, which grows optimally from 55 °C to 65 °C and is the main cause of spoilage of low-acid canned foods (LACFs) at high temperature. Resistance to wet-heat, biocides and UV-C of spores formed at different temperatures was assessed either for a selection of M. thermoacetica strains or for the strain M. thermoacetica ATCC 39073. Spores formed at 45 °C were significantly more sensitive to wet-heat than spores produced at 55 °C, while spores produced at 65 °C were as heat-resistant as spores produced at 55 °C. Spores of M. thermoacetica ATCC 39073 produced at 45 °C were significantly less resistant to peracetic acid than spores formed at 55 °C, while no difference in sensitivity to H2O2 or to UV-C treatment was observed whatever the sporulation temperature. However, both types of treatment enabled at least a 3.3 log CFU/mL reduction of M. thermoacetica ATCC 39073 spores. M. thermoacetica spores thus showed higher resistance properties when sporulation temperature was close to optimal growth temperature. These findings suggest food spoilage due to M. thermoacetica species could be controllable by holding temperatures below optimal growth temperature from the blanching step to the can filling step.


Assuntos
Moorella/crescimento & desenvolvimento , Esporos Bacterianos/química , Alimentos em Conserva/microbiologia , Temperatura Alta , Peróxido de Hidrogênio/farmacologia , Moorella/química , Moorella/efeitos dos fármacos , Ácido Peracético/farmacologia , Esporos Bacterianos/efeitos dos fármacos , Esporos Bacterianos/crescimento & desenvolvimento , Temperatura
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