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Nature Structural & Molecular Biology Apr 2023Alkane monooxygenase (AlkB) is a widely occurring integral membrane metalloenzyme that catalyzes the initial step in the functionalization of recalcitrant alkanes with...
Alkane monooxygenase (AlkB) is a widely occurring integral membrane metalloenzyme that catalyzes the initial step in the functionalization of recalcitrant alkanes with high terminal selectivity. AlkB enables diverse microorganisms to use alkanes as their sole carbon and energy source. Here we present the 48.6-kDa cryo-electron microscopy structure of a natural fusion from Fontimonas thermophila between AlkB and its electron donor AlkG at 2.76 Å resolution. The AlkB portion contains six transmembrane helices with an alkane entry tunnel within its transmembrane domain. A dodecane substrate is oriented by hydrophobic tunnel-lining residues to present a terminal C-H bond toward a diiron active site. AlkG, an [Fe-4S] rubredoxin, docks via electrostatic interactions and sequentially transfers electrons to the diiron center. The archetypal structural complex presented reveals the basis for terminal C-H selectivity and functionalization within this broadly distributed evolutionary class of enzymes.
Topics: Cryoelectron Microscopy; Cytochrome P-450 CYP4A; Alkanes
PubMed: 36997762
DOI: 10.1038/s41594-023-00958-0 -
Environmental Science and Pollution... Jun 2022Tobacco smoke (TS) is the source of a number of toxicants affecting the atmosphere and poses a threat to smokers and the whole community. Chemical, physical, and...
Tobacco smoke (TS) is the source of a number of toxicants affecting the atmosphere and poses a threat to smokers and the whole community. Chemical, physical, and toxicological features of smoking products (vapors as well as mainstream, side stream, and third-hand smoke) have been investigated extensively. Special attention is paid to organic compounds (individually or in combination giving rise to peculiar molecular fingerprints), potentially able to act as "chemical signature" of TS. In this regard, the percent distribution of long-chainnormal, iso, and anteiso alkanes was ascertained as typical of TS. Nevertheless, until now no indexes have been identified as suitable for assessing the global TS contribution to environmental pollution, e.g., the TS percentage in carbonaceous aerosol and in deposited dusts, the only exception consisting in the use of nicotelline as tracer. This paper describes the results of an extensive study aimed at chemically characterizing the nonpolar lipid fraction associated to suspended particulates (PMs) and deposition dusts (DDs) collected at indoor and outdoor locations. Based on the iso, anteiso, and normal C-C alkane profile in the samples as well in tobacco smoke- and no-TS-related emissions (literature data), various parameters describing the distribution of compounds were investigated. Finally, a cumulative variable was identified as the tobacco smoke impact index (TS%) suitable for estimating the TS percentage occurring in the particulate matter. The TS% rates were plotted vs. the exceedance of normal C alkane with respect to the average of C and C homologs, which results higher in TS than in most other emissions, revealing a link in the case of suspended particulates but not of deposited dusts. According to back analysis carried out on all particulate matter sets, it was found that traces of TS affect even remote areas, while inside the smokers' homes the contributions of TS to PM could account for up to ~61% and ~10%, respectively, in PM and DD. This confirms the need of valuing the health risk posed by TS to humans, by means of tools easy to apply in extensive investigations.
Topics: Air Pollution, Indoor; Alkanes; Dust; Environmental Monitoring; Humans; Particulate Matter; Tobacco Smoke Pollution
PubMed: 35092585
DOI: 10.1007/s11356-021-16617-0 -
Nature Microbiology Jul 2023Methanogenic and methanotrophic archaea produce and consume the greenhouse gas methane, respectively, using the reversible enzyme methyl-coenzyme M reductase (Mcr)....
Methanogenic and methanotrophic archaea produce and consume the greenhouse gas methane, respectively, using the reversible enzyme methyl-coenzyme M reductase (Mcr). Recently, Mcr variants that can activate multicarbon alkanes have been recovered from archaeal enrichment cultures. These enzymes, called alkyl-coenzyme M reductase (Acrs), are widespread in the environment but remain poorly understood. Here we produced anoxic cultures degrading mid-chain petroleum n-alkanes between pentane (C) and tetradecane (C) at 70 °C using oil-rich Guaymas Basin sediments. In these cultures, archaea of the genus Candidatus Alkanophaga activate the alkanes with Acrs and completely oxidize the alkyl groups to CO. Ca. Alkanophaga form a deep-branching sister clade to the methanotrophs ANME-1 and are closely related to the short-chain alkane oxidizers Ca. Syntrophoarchaeum. Incapable of sulfate reduction, Ca. Alkanophaga shuttle electrons released from alkane oxidation to the sulfate-reducing Ca. Thermodesulfobacterium syntrophicum. These syntrophic consortia are potential key players in petroleum degradation in heated oil reservoirs.
Topics: Archaea; Petroleum; Hydrothermal Vents; Anaerobiosis; Alkanes; Sulfates
PubMed: 37264141
DOI: 10.1038/s41564-023-01400-3 -
Microbial Biotechnology Jan 2017Bio-jet fuel has attracted a lot of interest in recent years and has become a focus for aircraft and engine manufacturers, oil companies, governments and researchers.... (Review)
Review
Bio-jet fuel has attracted a lot of interest in recent years and has become a focus for aircraft and engine manufacturers, oil companies, governments and researchers. Given the global concern about environmental issues and the instability of oil market, bio-jet fuel has been identified as a promising way to reduce the greenhouse gas emissions from the aviation industry, while also promoting energy security. Although a number of bio-jet fuel sources have been approved for manufacture, their commercialization and entry into the market is still a far way away. In this review, we provide an overview of the drivers for intensified research into bio-jet fuel technologies, the type of chemical compounds found in bio-jet fuel preparations and the current state of related pre-commercial technologies. The biosynthesis of hydrocarbons is one of the most promising approaches for bio-jet fuel production, and thus we provide a detailed analysis of recent advances in the microbial biosynthesis of hydrocarbons (with a focus on alkanes). Finally, we explore the latest developments and their implications for the future of research into bio-jet fuel technologies.
Topics: Alkanes; Bacteria; Biofuels; Biotechnology; Fungi
PubMed: 27723249
DOI: 10.1111/1751-7915.12423 -
Plant Physiology Oct 2022Cuticular waxes cover the aerial surfaces of land plants and protect them from various environmental stresses. Alkanes are major wax components and contribute to plant...
Cuticular waxes cover the aerial surfaces of land plants and protect them from various environmental stresses. Alkanes are major wax components and contribute to plant drought tolerance, but the biosynthesis and regulation of alkanes remain largely unknown in wheat (Triticum aestivum L.). Here, we identified and functionally characterized a key alkane biosynthesis gene ECERIFERUM1-6A (TaCER1-6A) from wheat. The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9)-mediated knockout mutation in TaCER1-6A greatly reduced the contents of C27, C29, C31, and C33 alkanes in wheat leaves, while TaCER1-6A overexpression significantly increased the contents of these alkanes in wheat leaves, suggesting that TaCER1-6A is specifically involved in the biosynthesis of C27, C29, C31, and C33 alkanes on wheat leaf surfaces. TaCER1-6A knockout lines exhibited increased cuticle permeability and reduced drought tolerance, whereas TaCER1-6A overexpression lines displayed reduced cuticle permeability and enhanced drought tolerance. TaCER1-6A was highly expressed in flag leaf blades and seedling leaf blades and could respond to abiotic stresses and abscisic acid. TaCER1-6A was located in the endoplasmic reticulum, which is the subcellular compartment responsible for wax biosynthesis. A total of three haplotypes (HapI/II/III) of TaCER1-6A were identified in 43 wheat accessions, and HapI was the dominant haplotype (95%) in these wheat varieties. Additionally, we identified two R2R3-MYB transcription factors TaMYB96-2D and TaMYB96-5D that bound directly to the conserved motif CAACCA in promoters of the cuticular wax biosynthesis genes TaCER1-6A, TaCER1-1A, and fatty acyl-CoA reductase4. Collectively, these results suggest that TaCER1-6A is required for C27, C29, C31, and C33 alkanes biosynthesis and improves drought tolerance in wheat.
Topics: Triticum; Droughts; Alkanes; Gene Expression Regulation, Plant; Waxes; Plant Leaves
PubMed: 36000923
DOI: 10.1093/plphys/kiac394 -
The Journal of Biological Chemistry Dec 1987The OCT plasmid encodes enzymes for alkane hydroxylation and alkanol dehydrogenation. Structural components are encoded on the 7.5-kilobase pair alkBAC operon, whereas...
The OCT plasmid encodes enzymes for alkane hydroxylation and alkanol dehydrogenation. Structural components are encoded on the 7.5-kilobase pair alkBAC operon, whereas positive regulatory components are encoded by alkR. We have constructed plasmids containing fusions of cloned alkBAC and alkR DNA and used these fusion plasmids to study the functional expression of the alkBAC operon and the regulatory locus alkR in Pseudomonas putida and in Escherichia coli. Growth on alkanes requires a functional chromosomally encoded fatty acid degradation system in addition to the plasmid-borne alk system. While such a system is active in P. putida, it is active in E. coli only in fadR mutants in which fatty acid degradation enzymes are expressed constitutively. Using such mutants, we found that E. coli as well as P. putida grew on octane as the sole source of carbon and energy when they were supplied with the cloned complete alk system. The alkR locus was strictly necessary in E. coli as well as in P. putida for expression of the alkBAC operon. The alkBAC operon could, however, be further reduced to a 5-kilobase pair operon without affecting the Alk phenotype in either species to a significant extent. Although with this reduction the plasmid-encoded alkanol dehydrogenase activity was lost, chromosomally encoded alkanol dehydrogenases in P. putida and E. coli compensated for this loss. The induction kinetics of the alk system was studied in detail in P. putida and E. coli. We used specific antibodies raised against alkane hydroxylase to follow the appearance of this protein following induction with octane. We found the induction kinetics of alkane hydroxylase to be similar in both species. A steady-state level was reached after about 2 h of induction in which time the alkane hydroxylase accounted for about 1.5% of total newly synthesized protein. Thus, alkBAC expression is very efficient and strictly regulated to both P. putida and E. coli.
Topics: Alkanes; Cloning, Molecular; Cytochrome P-450 CYP4A; Cytochrome P-450 Enzyme System; DNA Restriction Enzymes; Escherichia coli; Genotype; Kinetics; Mixed Function Oxygenases; Octanes; Operon; Phenotype; Plasmids; Pseudomonas
PubMed: 2826430
DOI: No ID Found -
Applied and Environmental Microbiology Feb 2019In subduction zones, serpentinization and biological processes may release alkanes to the deep waters, which would probably result in the rapid spread of However, the...
In subduction zones, serpentinization and biological processes may release alkanes to the deep waters, which would probably result in the rapid spread of However, the timing and area of the alkane distribution and associated enrichment of alkane-degrading microbes in the dark world of the deep ocean have not been explored. In this study, we report the richness (up to 17.8%) of alkane-degrading bacteria, represented by , in deep water samples obtained at 3,000 to 6,000 m in the Mariana Trench in two cruises. The relative abundance of correlated with copy numbers of functional and genes, which are involved in alkane degradation. In these water samples, we detected a high flux of alkanes, which probably resulted in the prevalence of in the deep waters. Contigs of were binned from the metagenomes for examination of alkane degradation pathways and deep sea-specific pathways, which revealed a lack of nitrate and nitrite dissimilatory reduction in our strains. Comparing the results for the two cruises conducted close to each other, we suggest periodic release of alkanes that may spread widely but periodically in the trench. Distribution of alkane-degrading bacteria in the world's oceans suggests the periodic and remarkable contributions of to the deep sea organic carbon and nitrogen sources. In the oligotrophic environment of the Mariana Trench, alkanes as carbohydrates are important for the ecosystem, but their spatial and periodic spreading in deep waters has never been reported. Alkane-degrading bacteria such as spp. are biological signals of the alkane distribution. In the present study, was abundant in some waters, at depths of up to 6,000 m, in the Mariana Trench. Genomic, transcriptomic, and chemical analyses provide evidence for the presence and activities of in deep sea zones. The periodic spreading of alkanes, probably from the subductive plates, might have fundamentally modified the local microbial communities, as well as perhaps the deep sea microenvironment.
Topics: Alcanivoraceae; Alkanes; Biodegradation, Environmental; Ecosystem; Nitrates; Nitrites; Phylogeny; Seawater
PubMed: 30446553
DOI: 10.1128/AEM.02089-18 -
Journal of Bacteriology Mar 1980The cellular structure of two yeast strains capable of growth on methane was investigated by electron microscopy. Microbodies were observed in cells of Sporobolomyces...
The cellular structure of two yeast strains capable of growth on methane was investigated by electron microscopy. Microbodies were observed in cells of Sporobolomyces roseus strain Y and Rhodotorula glutinis strain CY when grown on methane but rarely when grown on glucose. The size of the microbodies and the number observed per cell in a thin section did not increase with culture age. No crystalline organization was observed within these organelles. Similar microbodies were also observed in cells of R. glutinis CY grown on hexadecane. The plasma membranes of both methane and hexadecane-grown cells exhibited increased invagination compared to that of glucose-grown cells. Catalase activity was detected in the microbodies of alkane-grown cells by using 3,3'-diaminobenzidine as a cytochemical stain. The data presented suggest that microbodies, and the catalase contained within them, play a role in eucaryotic methane metabolism.
Topics: Alkanes; Catalase; Cell Membrane; Glucose; Methane; Microbodies; Microscopy, Electron; Mitosporic Fungi; Organoids; Rhodotorula
PubMed: 7189193
DOI: 10.1128/jb.141.3.1340-1349.1980 -
Nature Communications Dec 2014Alkane-degrading bacteria are ubiquitous in marine environments, but little is known about how alkane degradation is regulated. Here we investigate alkane sensing,...
Alkane-degrading bacteria are ubiquitous in marine environments, but little is known about how alkane degradation is regulated. Here we investigate alkane sensing, chemotaxis, signal transduction, uptake and pathway regulation in Alcanivorax dieselolei. The outer membrane protein OmpS detects the presence of alkanes and triggers the expression of an alkane chemotaxis complex. The coupling protein CheW2 of the chemotaxis complex, which is induced only by long-chain (LC) alkanes, sends signals to trigger the expression of Cyo, which participates in modulating the expression of the negative regulator protein AlmR. This change in turn leads to the expression of ompT1 and almA, which drive the selective uptake and hydroxylation of LC alkanes, respectively. AlmA is confirmed as a hydroxylase of LC alkanes. Additional factors responsible for the metabolism of medium-chain-length alkanes are also identified, including CheW1, OmpT1 and OmpT2. These results provide new insights into alkane metabolism pathways from alkane sensing to degradation.
Topics: Alcanivoraceae; Alkanes; Bacterial Outer Membrane Proteins; Biodegradation, Environmental; Cell Membrane; Chemotaxis; Gene Expression Regulation, Bacterial; Gene Library; Gene Regulatory Networks; Hydroxylation; Metabolic Networks and Pathways; Seawater; Signal Transduction
PubMed: 25502912
DOI: 10.1038/ncomms6755 -
Genomics Jan 2021Genome of Alcanivorax sp. IO_7, an alkane degrading deep-sea bacteria isolated from hydrothermally-influenced Southwest Indian Ridge was sequenced and analysed. Genomic...
Complete genome sequence and comparative genome analysis of Alcanivorax sp. IO_7, a marine alkane-degrading bacterium isolated from hydrothermally-influenced deep seawater of southwest Indian ridge.
Genome of Alcanivorax sp. IO_7, an alkane degrading deep-sea bacteria isolated from hydrothermally-influenced Southwest Indian Ridge was sequenced and analysed. Genomic data mining revealed gene clusters for degrading n-alkane and cycloalkanes, including biosurfactant production. The strain was shown to grow on hexadecane as its sole carbon source, supporting the findings of genomic analysis. Presence of cyclohexanone monooxygenase among genomic islands suggest that this strain may have used gene transfer to enhance its hydrocarbon degradation ability. Genes encoding for heavy metal resistance, multidrug resistance and multiple natural product biosynthesis crucial for survival in the hydrothermally influenced deep sea environment were detected. In our comparative genome analysis, it was evident that marine Alcanivorax strains contain a suite of enzymes for n-alkane and haloalkanoate degradation. Comparative genome and genomic synteny analysis provided insights into the physiological features and adaptation strategies of Alcanivorax sp. IO_7 in the deep-sea hydrothermal environment.
Topics: Alcanivoraceae; Alkanes; Drug Resistance; Genome, Bacterial; Genomic Islands; Metals, Heavy; Seawater
PubMed: 33096255
DOI: 10.1016/j.ygeno.2020.10.020