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Biology Nov 2023Iron is an essential nutrient for all life forms. Specialized mechanisms exist in bacteria to ensure iron uptake and its delivery to key enzymes within the cell, while... (Review)
Review
Iron is an essential nutrient for all life forms. Specialized mechanisms exist in bacteria to ensure iron uptake and its delivery to key enzymes within the cell, while preventing toxicity. Iron uptake and exchange networks must adapt to the different environmental conditions, particularly those that require the biosynthesis of multiple iron proteins, such as nitrogen fixation. In this review, we outline the mechanisms that the model diazotrophic bacterium uses to ensure iron nutrition and how it adapts Fe metabolism to diazotrophic growth.
PubMed: 37998022
DOI: 10.3390/biology12111423 -
Journal of Biological Inorganic... Mar 2015Nitrogenase catalyzes biological nitrogen fixation, a key step in the global nitrogen cycle. Three homologous nitrogenases have been identified to date, along with... (Review)
Review
Nitrogenase catalyzes biological nitrogen fixation, a key step in the global nitrogen cycle. Three homologous nitrogenases have been identified to date, along with several structural and/or functional homologs of this enzyme that are involved in nitrogenase assembly, bacteriochlorophyll biosynthesis and methanogenic process, respectively. In this article, we provide an overview of the structures and functions of nitrogenase and its homologs, which highlights the similarity and disparity of this uniquely versatile group of enzymes.
Topics: Azotobacter vinelandii; Bacteriochlorophylls; Catalysis; Molybdenum; Nitrogen; Nitrogen Fixation; Nitrogenase; Structure-Activity Relationship
PubMed: 25491285
DOI: 10.1007/s00775-014-1225-3 -
Microbiology (Reading, England) Apr 2018Azotobacter vinelandii has been studied for over 100 years since its discovery as an aerobic nitrogen-fixing organism. This species has proved useful for the study of... (Review)
Review
Azotobacter vinelandii has been studied for over 100 years since its discovery as an aerobic nitrogen-fixing organism. This species has proved useful for the study of many different biological systems, including enzyme kinetics and the genetic code. It has been especially useful in working out the structures and mechanisms of different nitrogenase enzymes, how they can function in oxic environments and the interactions of nitrogen fixation with other aspects of metabolism. Interest in studying A. vinelandii has waned in recent decades, but this bacterium still possesses great potential for new discoveries in many fields and commercial applications. The species is of interest for research because of its genetic pliability and natural competence. Its features of particular interest to industry are its ability to produce multiple valuable polymers - bioplastic and alginate in particular; its nitrogen-fixing prowess, which could reduce the need for synthetic fertilizer in agriculture and industrial fermentations, via coculture; its production of potentially useful enzymes and metabolic pathways; and even its biofuel production abilities. This review summarizes the history and potential for future research using this versatile microbe.
Topics: Azotobacter vinelandii; Biofuels; Biopolymers; Hydrogen; Metabolic Engineering; Metabolic Networks and Pathways; Nitrogen; Nitrogenase; Oxidoreductases; Oxygen
PubMed: 29533747
DOI: 10.1099/mic.0.000643 -
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 -
Biology Letters Mar 2022Declining food production in African agroecosystems is attributable to changes in weather patterns, soil infertility and limited farming inputs. The exploitation of...
Declining food production in African agroecosystems is attributable to changes in weather patterns, soil infertility and limited farming inputs. The exploitation of plant growth-promoting soil microbes could remedy these problems. Such microbes include ; free-living, nitrogen-fixing bacteria, which confer stress tolerance, avail phytohormones and aid in soil bioremediation. Here, we aimed to isolate, characterize and determine the biodiversity of native isolates from soils in semi-arid Eastern Kenya. Isolation was conducted on nitrogen-free Ashby's agar and the morphological, biochemical and molecular attributes evaluated. The isolates were sequenced using DNA amplicons of 27F and 1492R primers of the 16S rRNA gene loci. The Basic Local Alignment Search Tool (BLASTn) analysis of their sequences revealed the presence of three main species viz., and . Kitui County recorded the highest number of recovered isolates (45.4%) and lowest diversity index (0.8761). Tharaka Nithi County showed the lowest occurrence (26.36%) with a diversity index of (1.057). The diversity was influenced by the soil pH, texture and total organic content. This study reports for the first time a wide diversity of species from a semi-arid agroecosystem in Kenya with potential for utilization as low-cost, free-living nitrogen-fixing bioinoculant.
Topics: Azotobacter; Kenya; Nitrogen; RNA, Ribosomal, 16S; Soil; Soil Microbiology
PubMed: 35317624
DOI: 10.1098/rsbl.2021.0612 -
Frontiers in Microbiology 2022Alginates are a family of polymers composed of guluronate and mannuronate monomers joined by β (1-4) links. The different types of alginates have variations in their... (Review)
Review
Alginates are a family of polymers composed of guluronate and mannuronate monomers joined by β (1-4) links. The different types of alginates have variations in their monomer content and molecular weight, which determine the rheological properties and their applications. In industry, alginates are commonly used as additives capable of viscosifying, stabilizing, emulsifying, and gelling aqueous solutions. Recently, additional specialized biomedical uses have been reported for this polymer. Currently, the production of alginates is based on the harvesting of seaweeds; however, the composition and structure of the extracts are highly variable. The production of alginates for specialized applications requires a precise composition of monomers and molecular weight, which could be achieved using bacterial production systems such as those based on , a free-living, non-pathogenic bacterium. In this mini-review, we analyze the latest advances in the regulation of alginate synthesis in this model.
PubMed: 35401471
DOI: 10.3389/fmicb.2022.845473 -
Protein Science : a Publication of the... Oct 2017Azotobacter vinelandii flavodoxin II serves as a physiological reductant of nitrogenase, the enzyme system mediating biological nitrogen fixation. Wildtype A. vinelandii...
Azotobacter vinelandii flavodoxin II serves as a physiological reductant of nitrogenase, the enzyme system mediating biological nitrogen fixation. Wildtype A. vinelandii flavodoxin II was electrochemically and crystallographically characterized to better understand the molecular basis for this functional role. The redox properties were monitored on surfactant-modified basal plane graphite electrodes, with two distinct redox couples measured by cyclic voltammetry corresponding to reduction potentials of -483 ± 1 mV and -187 ± 9 mV (vs. NHE) in 50 mM potassium phosphate, 150 mM NaCl, pH 7.5. These redox potentials were assigned as the semiquinone/hydroquinone couple and the quinone/semiquinone couple, respectively. This study constitutes one of the first applications of surfactant-modified basal plane graphite electrodes to characterize the redox properties of a flavodoxin, thus providing a novel electrochemical method to study this class of protein. The X-ray crystal structure of the flavodoxin purified from A. vinelandii was solved at 1.17 Å resolution. With this structure, the native nitrogenase electron transfer proteins have all been structurally characterized. Docking studies indicate that a common binding site surrounding the Fe-protein [4Fe:4S] cluster mediates complex formation with the redox partners Mo-Fe protein, ferredoxin I, and flavodoxin II. This model supports a mechanistic hypothesis that electron transfer reactions between the Fe-protein and its redox partners are mutually exclusive.
Topics: Azotobacter vinelandii; Bacterial Proteins; Crystallography, X-Ray; Electrochemistry; Flavodoxin; Hydrogen-Ion Concentration; Iron; Models, Molecular; Nitrogenase; Protein Conformation
PubMed: 28710816
DOI: 10.1002/pro.3236 -
Antioxidants & Redox Signaling Feb 2020Structural and functional characterization of the globin-coupled sensors (GCSs) from (GReg) and (GReg). Ultraviolet/visible and resonance Raman spectroscopies...
Structural and functional characterization of the globin-coupled sensors (GCSs) from (GReg) and (GReg). Ultraviolet/visible and resonance Raman spectroscopies confirm the presence in GReg and GReg of a globin domain capable of reversible gaseous ligand binding. In GReg, an influence of the transmitter domain on the heme proximal region of the globin domain can be seen, and ' is higher than for other GCSs. The O binding kinetics suggests the presence of an open and a closed conformation. As for GReg, the fully oxygenated GReg show a very high diguanylate cyclase activity. The carbon monoxide rebinding to GReg indicates that intra- and intermolecular interactions influence the ligand binding. The globin domains of both proteins (GReg globin domain and GRegGb with cysteines (Cys16, 45, 114, 154) mutated to serines [GReg-Gb*]) share the same GCS fold, a similar proximal but a different distal side structure. They homodimerize through a G-H helical bundle as in other GCSs. However, GReg-Gb* shows also a second dimerization mode. This article extends our knowledge on the GCS proteins and contributes to a better understanding of the GCSs role in the formation of bacterial biofilms. GReg and GReg conform to the GCS family, share a similar overall structure, but they have different properties in terms of the ligand binding. In particular, GReg shows an open and a closed conformation that in the latter form will very tightly bind oxygen. GReg has only one closed conformation. In both proteins, it is the fully oxygenated GCS form that catalyzes the production of the second messenger.
Topics: Azotobacter vinelandii; Bacterial Proteins; Binding Sites; Bordetella pertussis; Globins; Heme-Binding Proteins; Protein Structure, Quaternary; Protein Structure, Tertiary; Structure-Activity Relationship
PubMed: 31559835
DOI: 10.1089/ars.2018.7690 -
Journal of Bacteriology Nov 2021Azotobacter vinelandii is a nitrogen-fixing free-living soil microbe that has been studied for decades in relation to biological nitrogen fixation (BNF). It is highly...
Azotobacter vinelandii is a nitrogen-fixing free-living soil microbe that has been studied for decades in relation to biological nitrogen fixation (BNF). It is highly amenable to genetic manipulation, helping to unravel the intricate importance of different proteins involved in the process of BNF, including the biosynthesis of cofactors that are essential to assembling the complex metal cofactors that catalyze the difficult reaction of nitrogen fixation. Additionally, A. vinelandii accomplishes this feat while growing as an obligate aerobe, differentiating it from many of the nitrogen-fixing bacteria that are associated with plant roots. The ability to function in the presence of oxygen makes A. vinelandii suitable for application in various potential biotechnological schemes. In this study, we employed transposon sequencing (Tn-seq) to measure the fitness defects associated with disruptions of various genes under nitrogen-fixing dependent growth, versus growth with extraneously provided urea as a nitrogen source. The results allowed us to probe the importance of more than 3,800 genes, revealing that many genes previously believed to be important, can be successfully disrupted without impacting cellular fitness. These results provide insights into the functional redundancy in A. vinelandii, while also providing a direct measure of fitness for specific genes associated with the process of BNF. These results will serve as a valuable reference tool in future studies to uncover the mechanisms that govern this process.
Topics: Azotobacter vinelandii; Bacterial Proteins; Base Sequence; DNA Transposable Elements; Gene Expression Regulation, Bacterial; Genetic Fitness; Molybdenum; Nitrogen; Urea
PubMed: 34570624
DOI: 10.1128/JB.00404-21 -
Applied and Environmental Microbiology Sep 2022Biological nitrogen fixation requires large amounts of energy in the form of ATP and low potential electrons to overcome the high activation barrier for cleavage of the...
Biological nitrogen fixation requires large amounts of energy in the form of ATP and low potential electrons to overcome the high activation barrier for cleavage of the dinitrogen triple bond. The model aerobic nitrogen-fixing bacteria, Azotobacter vinelandii, generates low potential electrons in the form of reduced ferredoxin (Fd) and flavodoxin (Fld) using two distinct mechanisms via the enzyme complexes Rnf and Fix. Both Rnf and Fix are expressed during nitrogen fixation, but deleting either or genes has little effect on diazotrophic growth. However, deleting both and eliminates the ability to grow diazotrophically. Rnf and Fix both use NADH as a source of electrons, but overcoming the energetics of NADH's endergonic reduction of Fd/Fld is accomplished through different mechanisms. Rnf harnesses free energy from the chemiosmotic potential, whereas Fix uses electron bifurcation to effectively couple the endergonic reduction of Fd/Fld to the exergonic reduction of quinone. Different reaction stoichiometries and condition-specific differential gene expression indicate specific roles for the two reactions. This work's complementary physiological studies and thermodynamic modeling reveal how Rnf and Fix balance redox homeostasis in various conditions. Specifically, the Fix complex is required for efficient growth under low oxygen concentrations, while Rnf is presumed to maintain reduced Fd/Fld production for nitrogenase under standard conditions. This work provides a framework for understanding how the production of low potential electrons sustains robust nitrogen fixation in various conditions. The availability of fixed nitrogen is critical for life in many ecosystems, from extreme environments to agriculture. Due to the energy demands of biological nitrogen fixation, organisms must tailor their metabolism during diazotrophic growth to deliver the energy requirements to nitrogenase in the form of ATP and low potential electrons. Therefore, a complete understanding of diazotrophic energy metabolism and redox homeostasis is required to understand the impact on ecological communities or to promote crop growth in agriculture through engineered diazotrophs.
Topics: Adenosine Triphosphate; Azotobacter vinelandii; Ecosystem; Ferredoxins; NAD; Nitrogen; Nitrogen Fixation; Nitrogenase
PubMed: 36000884
DOI: 10.1128/aem.01049-22