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The FEBS Journal Oct 2017The flavodoxin-like fold is a protein architecture that can be traced back to the universal ancestor of the three kingdoms of life. Many proteins share this α-β... (Review)
Review
The flavodoxin-like fold is a protein architecture that can be traced back to the universal ancestor of the three kingdoms of life. Many proteins share this α-β parallel topology and hence it is highly relevant to illuminate how they fold. Here, we review experiments and simulations concerning the folding of flavodoxins and CheY-like proteins, which share the flavodoxin-like fold. These polypeptides tend to temporarily misfold during unassisted folding to their functionally active forms. This susceptibility to frustration is caused by the more rapid formation of an α-helix compared to a β-sheet, particularly when a parallel β-sheet is involved. As a result, flavodoxin-like proteins form intermediates that are off-pathway to native protein and several of these species are molten globules (MGs). Experiments suggest that the off-pathway species are of helical nature and that flavodoxin-like proteins have a nonconserved transition state that determines the rate of productive folding. Folding of flavodoxin from Azotobacter vinelandii has been investigated extensively, enabling a schematic construction of its folding energy landscape. It is the only flavodoxin-like protein of which cotranslational folding has been probed. New insights that emphasize differences between in vivo and in vitro folding energy landscapes are emerging: the ribosome modulates MG formation in nascent apoflavodoxin and forces this polypeptide toward the native state.
Topics: Azotobacter vinelandii; Escherichia coli; Escherichia coli Proteins; Flavodoxin; Gene Expression; Methyl-Accepting Chemotaxis Proteins; Models, Molecular; Protein Biosynthesis; Protein Conformation, alpha-Helical; Protein Conformation, beta-Strand; Protein Folding; Protein Isoforms; Thermodynamics
PubMed: 28380286
DOI: 10.1111/febs.14077 -
Biomolecules Aug 2022Flavodoxins are small electron transport proteins that are involved in a myriad of photosynthetic and non-photosynthetic metabolic pathways in Bacteria (including...
Flavodoxins are small electron transport proteins that are involved in a myriad of photosynthetic and non-photosynthetic metabolic pathways in Bacteria (including cyanobacteria), Archaea and some algae. The sequenced genome of 0305φ8-36, a large bacteriophage that infects the soil bacterium , was predicted to encode a putative flavodoxin redox protein. Here we confirm that 0305φ8-36 phage encodes a FMN-containing flavodoxin polypeptide and we report the expression, purification and enzymatic characterization of the recombinant protein. Purified 0305φ8-36 flavodoxin has near-identical spectral properties to control, purified flavodoxin. Using in vitro assays we show that 0305φ8-36 flavodoxin can be reconstituted with flavodoxin reductase and support regio- and stereospecific cytochrome P450 CYP170A1 allyl-oxidation of epi-isozizaene to the sesquiterpene antibiotic product albaflavenone, found in the soil bacterium . In vivo, 0305φ8-36 flavodoxin is predicted to mediate the 2-electron reduction of the β subunit of phage-encoded ribonucleotide reductase to catalyse the conversion of ribonucleotides to deoxyribonucleotides during viral replication. Our results demonstrate that this phage flavodoxin has the potential to manipulate and drive bacterial P450 cellular metabolism, which may affect both the host biological fitness and the communal microbiome. Such a scenario may also be applicable in other viral-host symbiotic/parasitic relationships.
Topics: Cytochrome P-450 Enzyme System; Escherichia coli; Flavodoxin; Oxidation-Reduction; Soil; Streptomyces coelicolor
PubMed: 36009001
DOI: 10.3390/biom12081107 -
Journal of Experimental Botany Oct 2014Ferredoxins are electron shuttles harbouring iron-sulfur clusters that connect multiple oxido-reductive pathways in organisms displaying different lifestyles. Some... (Review)
Review
Ferredoxins are electron shuttles harbouring iron-sulfur clusters that connect multiple oxido-reductive pathways in organisms displaying different lifestyles. Some prokaryotes and algae express an isofunctional electron carrier, flavodoxin, which contains flavin mononucleotide as cofactor. Both proteins evolved in the anaerobic environment preceding the appearance of oxygenic photosynthesis. The advent of an oxygen-rich atmosphere proved detrimental to ferredoxin owing to iron limitation and oxidative damage to the iron-sulfur cluster, and many microorganisms induced flavodoxin expression to replace ferredoxin under stress conditions. Paradoxically, ferredoxin was maintained throughout the tree of life, whereas flavodoxin is absent from plants and animals. Of note is that flavodoxin expression in transgenic plants results in increased tolerance to multiple stresses and iron deficit, through mechanisms similar to those operating in microorganisms. Then, the question remains open as to why a trait that still confers plants such obvious adaptive benefits was not retained. We compare herein the properties of ferredoxin and flavodoxin, and their contrasting modes of expression in response to different environmental stimuli. Phylogenetic analyses suggest that the flavodoxin gene was already absent in the algal lineages immediately preceding land plants. Geographical distribution of phototrophs shows a bias against flavodoxin-containing organisms in iron-rich coastal/freshwater habitats. Based on these observations, we propose that plants evolved from freshwater macroalgae that already lacked flavodoxin because they thrived in an iron-rich habitat with no need to back up ferredoxin functions and therefore no selective pressure to keep the flavodoxin gene. Conversely, ferredoxin retention in the plant lineage is probably related to its higher efficiency as an electron carrier, compared with flavodoxin. Several lines of evidence supporting these contentions are presented and discussed.
Topics: Biological Evolution; Electron Transport; Ferredoxins; Flavodoxin; Iron; Phylogeny
PubMed: 25009172
DOI: 10.1093/jxb/eru273 -
Antioxidants (Basel, Switzerland) Oct 2022Distinct isoforms of FAD-containing ferredoxin-NADP oxidoreductase (FNR) and ferredoxin (Fd) are involved in photosynthetic and non-photosynthetic electron transfer... (Review)
Review
Distinct isoforms of FAD-containing ferredoxin-NADP oxidoreductase (FNR) and ferredoxin (Fd) are involved in photosynthetic and non-photosynthetic electron transfer systems. The FNR (FAD)-Fd [2Fe-2S] redox pair complex switches between one- and two-electron transfer reactions in steps involving FAD semiquinone intermediates. In cyanobacteria and some algae, one-electron carrier Fd serves as a substitute for low-potential FMN-containing flavodoxin (Fld) during growth under low-iron conditions. This complex evolves into the covalent FNR (FAD)-Fld (FMN) pair, which participates in a wide variety of NAD(P)H-dependent metabolic pathways as an electron donor, including bacterial sulfite reductase, cytochrome P450 BM3, plant or mammalian cytochrome P450 reductase and nitric oxide synthase isoforms. These electron transfer systems share the conserved Ser-Glu/Asp pair in the active site of the FAD module. In addition to physiological electron acceptors, the NAD(P)H-dependent diflavin reductase family catalyzes a one-electron reduction of artificial electron acceptors such as quinone-containing anticancer drugs. Conversely, NAD(P)H: quinone oxidoreductase (NQO1), which shares a Fld-like active site, functions as a typical two-electron transfer antioxidant enzyme, and the NQO1 and UDP-glucuronosyltransfease/sulfotransferase pairs function as an antioxidant detoxification system. In this review, the roles of the plant FNR-Fd and FNR-Fld complex pairs were compared to those of the diflavin reductase (FAD-FMN) family. In the final section, evolutionary aspects of NAD(P)H-dependent multi-domain electron transfer systems are discussed.
PubMed: 36358515
DOI: 10.3390/antiox11112143 -
Life (Basel, Switzerland) Mar 2015Flavodiiron proteins (FDPs, also called flavoproteins, Flvs) are modular enzymes widely present in Bacteria and Archaea. The evolution of cyanobacteria and oxygenic... (Review)
Review
Flavodiiron proteins (FDPs, also called flavoproteins, Flvs) are modular enzymes widely present in Bacteria and Archaea. The evolution of cyanobacteria and oxygenic photosynthesis occurred in concert with the modulation of typical bacterial FDPs. Present cyanobacterial FDPs are composed of three domains, the β-lactamase-like, flavodoxin-like and flavin-reductase like domains. Cyanobacterial FDPs function as hetero- and homodimers and are involved in the regulation of photosynthetic electron transport. Whilst Flv2 and Flv4 proteins are limited to specific cyanobacterial species (β-cyanobacteria) and function in photoprotection of Photosystem II, Flv1 and Flv3 proteins, functioning in the "Mehler-like" reaction and safeguarding Photosystem I under fluctuating light conditions, occur in nearly all cyanobacteria and additionally in green algae, mosses and lycophytes. Filamentous cyanobacteria have additional FDPs in heterocyst cells, ensuring a microaerobic environment for the function of the nitrogenase enzyme under the light. Here, the evolution, occurrence and functional mechanisms of various FDPs in oxygenic photosynthetic organisms are discussed.
PubMed: 25761262
DOI: 10.3390/life5010716 -
International Journal of Molecular... Mar 2020Flavodoxins are small soluble electron transfer proteins widely present in bacteria and absent in vertebrates. Flavodoxins participate in different metabolic pathways... (Review)
Review
Flavodoxins are small soluble electron transfer proteins widely present in bacteria and absent in vertebrates. Flavodoxins participate in different metabolic pathways and, in some bacteria, they have been shown to be essential proteins representing promising therapeutic targets to fight bacterial infections. Using purified flavodoxin and chemical libraries, leads can be identified that block flavodoxin function and act as bactericidal molecules, as it has been demonstrated for (), the most prevalent human gastric pathogen. Increasing antimicrobial resistance by this bacterium has led current therapies to lose effectiveness, so alternative treatments are urgently required. Here, we summarize, with a focus on flavodoxin, opportunities for pharmacological intervention offered by the potential protein targets described for this bacterium and provide information on other gastrointestinal pathogens and also on bacteria from the gut microbiota that contain flavodoxin. The process of discovery and development of novel antimicrobials specific for flavodoxin that is being carried out in our group is explained, as it can be extrapolated to the discovery of inhibitors specific for other gastric pathogens. The high specificity for of the antimicrobials developed may be of help to reduce damage to the gut microbiota and to slow down the development of resistant mutants.
Topics: Anti-Bacterial Agents; Bacterial Proteins; Drug Design; Drug Resistance, Bacterial; Flavodoxin; Gastrointestinal Microbiome; Helicobacter Infections; Helicobacter pylori; Humans; Molecular Targeted Therapy
PubMed: 32164177
DOI: 10.3390/ijms21051881 -
Microbiology Spectrum Oct 2021The strict anaerobe Clostridium ljungdahlii can ferment CO or H/CO via the Wood-Ljungdahl pathway to acetate, ethanol, and 2,3-butanediol. This ability has attracted...
The strict anaerobe Clostridium ljungdahlii can ferment CO or H/CO via the Wood-Ljungdahl pathway to acetate, ethanol, and 2,3-butanediol. This ability has attracted considerable interest, since it can be used for syngas fermentation to produce biofuels and biochemicals. However, the key enzyme methylenetetrahydrofolate reductase (MTHFR) in the Wood-Ljungdahl pathway of the strain has not been characterized, and its physiological electron donor is unclear. In this study, we purified the enzyme 46-fold with a benzyl viologen reduction activity of 41.2 U/mg from C. ljungdahlii cells grown on CO. It is composed of two subunits, MetF (31.5 kDa) and MetV (23.5 kDa), and has an apparent molecular mass of 62.2 kDa. The brownish yellow protein contains 0.73 flavin mononucleotide (FMN) and 7.4 Fe, in agreement with the prediction that MetF binds one flavin and MetV binds two [4Fe4S] clusters. It cannot use NAD(P)H as its electron donor or catalyze an electron-bifurcating reaction in combination with ferredoxin as an electron acceptor. The reduced recombinant ferredoxin, flavodoxin, and thioredoxin of can serve as electron donors with specific activities of 91.2, 22.1, and 7.4 U/mg, respectively. The apparent values for reduced ferredoxin and flavodoxin were around 1.46 μM and 0.73 μM, respectively. Subunit composition and phylogenetic analysis showed that the enzyme from belongs to MetFV-type MTHFR, which is a heterodimer, and uses reduced ferredoxin as its electron donor. Based on these results, we discuss the energy metabolism of when it grows on CO or H plus CO. Syngas, a mixture of CO, CO, and H, is the main component of steel mill waste gas and also can be generated by the gasification of biomass and urban domestic waste. Its fermentation to biofuels and biocommodities has attracted attention due to the economic and environmental benefits of this process. Clostridium ljungdahlii is one of the superior acetogens used in the technology. However, the biochemical mechanism of its gas fermentation via the Wood-Ljungdahl pathway is not completely clear. In this study, the key enzyme, methylenetetrahydrofolate reductase (MTHFR), was characterized and found to be a non-electron-bifurcating heterodimer with reduced ferredoxin as its electron donor, representing another example of MetFV-type MTHFR. The findings will form the basis for a deeper understanding of the energy metabolism of syngas fermentation by , which is valuable for developing metabolic engineering strains and efficient syngas fermentation technologies.
Topics: Biofuels; Carbon Dioxide; Carbon Monoxide; Clostridium; Energy Metabolism; Fermentation; Ferredoxins; Flavodoxin; Hydrogen; Methylenetetrahydrofolate Reductase (NADPH2); Thioredoxins
PubMed: 34643446
DOI: 10.1128/Spectrum.00958-21 -
Biochimica Et Biophysica Acta.... Apr 2024Reactions catalysed by iron-sulfur (Fe-S) enzymes appear in a variety of biosynthetic pathways that produce valuable natural products. Harnessing these biosynthetic... (Review)
Review
Reactions catalysed by iron-sulfur (Fe-S) enzymes appear in a variety of biosynthetic pathways that produce valuable natural products. Harnessing these biosynthetic pathways by expression in microbial cell factories grown on an industrial scale would yield enormous economic and environmental benefits. However, Fe-S enzymes often become bottlenecks that limits the productivity of engineered pathways. As a consequence, achieving the production metrics required for industrial application remains a distant goal for Fe-S enzyme-dependent pathways. Here, we identify and review three core challenges in harnessing Fe-S enzyme activity, which all stem from the properties of Fe-S clusters: 1) limited Fe-S cluster supply within the host cell, 2) Fe-S cluster instability, and 3) lack of specialized reducing cofactor proteins often required for Fe-S enzyme activity, such as enzyme-specific flavodoxins and ferredoxins. We highlight successful methods developed for a variety of Fe-S enzymes and electron carriers for overcoming these difficulties. We use heterologous nitrogenase expression as a grand case study demonstrating how each of these challenges can be addressed. We predict that recent breakthroughs in protein structure prediction and design will prove well-suited to addressing each of these challenges. A reliable toolkit for harnessing Fe-S enzymes in engineered metabolic pathways will accelerate the development of industry-ready Fe-S enzyme-dependent biosynthesis pathways.
PubMed: 38574823
DOI: 10.1016/j.bbamcr.2024.119718 -
Cyanobacterial hydrogenases and hydrogen metabolism revisited: recent progress and future prospects.International Journal of Molecular... May 2015Cyanobacteria have garnered interest as potential cell factories for hydrogen production. In conjunction with photosynthesis, these organisms can utilize inexpensive... (Review)
Review
Cyanobacteria have garnered interest as potential cell factories for hydrogen production. In conjunction with photosynthesis, these organisms can utilize inexpensive inorganic substrates and solar energy for simultaneous biosynthesis and hydrogen evolution. However, the hydrogen yield associated with these organisms remains far too low to compete with the existing chemical processes. Our limited understanding of the cellular hydrogen production pathway is a primary setback in the potential scale-up of this process. In this regard, the present review discusses the recent insight around ferredoxin/flavodoxin as the likely electron donor to the bidirectional Hox hydrogenase instead of the generally accepted NAD(P)H. This may have far reaching implications in powering solar driven hydrogen production. However, it is evident that a successful hydrogen-producing candidate would likely integrate enzymatic traits from different species. Engineering the [NiFe] hydrogenases for optimal catalytic efficiency or expression of a high turnover [FeFe] hydrogenase in these photo-autotrophs may facilitate the development of strains to reach target levels of biohydrogen production in cyanobacteria. The fundamental advancements achieved in these fields are also summarized in this review.
Topics: Cyanobacteria; Ferredoxins; Hydrogen; Hydrogenase; Iron-Sulfur Proteins; NADP
PubMed: 26006225
DOI: 10.3390/ijms160510537 -
Biochimica Et Biophysica Acta Oct 2001In plants and cyanobacteria, the primary step in oxygenic photosynthesis, the light induced charge separation, is driven by two large membrane intrinsic protein... (Review)
Review
In plants and cyanobacteria, the primary step in oxygenic photosynthesis, the light induced charge separation, is driven by two large membrane intrinsic protein complexes, the photosystems I and II. Photosystem I catalyses the light driven electron transfer from plastocyanin/cytochrome c(6) on the lumenal side of the membrane to ferredoxin/flavodoxin at the stromal side by a chain of electron carriers. Photosystem I of Synechococcus elongatus consists of 12 protein subunits, 96 chlorophyll a molecules, 22 carotenoids, three [4Fe4S] clusters and two phylloquinones. Furthermore, it has been discovered that four lipids are intrinsic components of photosystem I. Photosystem I exists as a trimer in the native membrane with a molecular mass of 1068 kDa for the whole complex. The X-ray structure of photosystem I at a resolution of 2.5 A shows the location of the individual subunits and cofactors and provides new information on the protein-cofactor interactions. [P. Jordan, P. Fromme, H.T. Witt, O. Klukas, W. Saenger, N. Krauss, Nature 411 (2001) 909-917]. In this review, biochemical data and results of biophysical investigations are discussed with respect to the X-ray crystallographic structure in order to give an overview of the structure and function of this large membrane protein.
Topics: Crystallography, X-Ray; Cyanobacteria; Light-Harvesting Protein Complexes; Membrane Proteins; Models, Chemical; Models, Molecular; Molecular Structure; Photosynthetic Reaction Center Complex Proteins; Photosystem I Protein Complex; Plant Proteins; Proteins
PubMed: 11687205
DOI: 10.1016/s0005-2728(01)00195-5