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FEMS Microbiology Ecology Oct 2013Alkane-degrading bacteria were isolated from uncontaminated soil microcosms, which had been incubated with maize litter as natural alkane source. The isolates served to... (Comparative Study)
Comparative Study
Alkane-degrading bacteria were isolated from uncontaminated soil microcosms, which had been incubated with maize litter as natural alkane source. The isolates served to understand spatio-temporal community changes at the soil-litter interface, which had been detected using alkB as a functional marker gene for bacterial alkane degraders. To obtain a large spectrum of isolates, liquid subcultivation was combined with a matrix-assisted enrichment (Teflon membranes, litter). Elevated cell numbers of alkane degraders were detected by most probable number counting indicating enhanced alkane degradation potential in soil in response to litter treatment. Partial 16S rRNA gene sequencing of 395 isolates revealed forty different phylogenetic groups [operational taxonomic units (OTUs)] and spatio-temporal shifts in community composition. Ten OTUs comprised so far unknown alkane degraders, and five OTUs represented putative new bacterial genera. The combination of enrichment methods yielded a higher diversity of isolates than liquid subcultivation alone. Comparison of 16S rRNA gene T-RFLP profiles indicated that many alkane degraders present in the enrichments were not detectable in the DNA extracts from soil microcosms. These possibly rare specialists might represent a seed bank for the alkane degradation capacity in uncontaminated soil. This relevant ecosystem function can be fostered by the formation of the soil-litter interface.
Topics: Alkanes; Bacteria; Ecosystem; Phylogeny; Polymorphism, Restriction Fragment Length; RNA, Ribosomal, 16S; Soil; Soil Microbiology
PubMed: 23406352
DOI: 10.1111/1574-6941.12097 -
Journal of Inorganic Biochemistry Jun 2021Interest in understanding the environmental distribution of the alkane monooxygenase (AlkB) enzyme led to the identification of over 100 distinct alkane monooxygenase...
Interest in understanding the environmental distribution of the alkane monooxygenase (AlkB) enzyme led to the identification of over 100 distinct alkane monooxygenase (AlkB) enzymes containing a covalently bound, or fused, rubredoxin. The rubredoxin-fused AlkB from Dietzia cinnamea was cloned as a full-length protein and as a truncated protein with the rubredoxin domain deleted. A point mutation (V91W) was introduced into the full-length protein, with the goal of assessing how steric bulk in the putative substrate channel might affect selectivity. Based on activity studies with alkane and alkene substrates, the rubredoxin-fused AlkB oxidizes a similar range of alkane substrates relative to its rubredoxin domain-deletion counterpart. Oxidation of terminal alkenes generated both an epoxide and a terminal aldehyde. The products of V91W-mutant-catalyzed oxidation of alkenes had a higher aldehyde-to-epoxide ratio than the products formed in the presence of the wild type protein. These results are consistent with this mutation causing a structural change impacting substrate positioning.
Topics: Actinobacteria; Alkanes; Alkenes; Bacterial Proteins; Catalysis; Computational Biology; Humans; Mixed Function Oxygenases; Oxidation-Reduction; Point Mutation; Prevalence; Rubredoxins
PubMed: 33752122
DOI: 10.1016/j.jinorgbio.2021.111409 -
BMC Genomics Dec 2014Psychrophiles are presumed to play a large role in the catabolism of alkanes and other components of crude oil in natural low temperature environments. In this study we...
BACKGROUND
Psychrophiles are presumed to play a large role in the catabolism of alkanes and other components of crude oil in natural low temperature environments. In this study we analyzed the functional diversity of genes for alkane hydroxylases, the enzymes responsible for converting alkanes to more labile alcohols, as found in the genomes of nineteen psychrophiles for which alkane degradation has not been reported. To identify possible mechanisms of low temperature optimization we compared putative alkane hydroxylases from these psychrophiles with homologues from nineteen taxonomically related mesophilic strains.
RESULTS
Seven of the analyzed psychrophile genomes contained a total of 27 candidate alkane hydroxylase genes, only two of which are currently annotated as alkane hydroxylase. These candidates were mostly related to the AlkB and cytochrome p450 alkane hydroxylases, but several homologues of the LadA and AlmA enzymes, significant for their ability to degrade long-chain alkanes, were also detected. These putative alkane hydroxylases showed significant differences in primary structure from their mesophile homologues, with preferences for specific amino acids and increased flexibility on loops, bends, and α-helices.
CONCLUSION
A focused analysis on psychrophile genomes led to discovery of numerous candidate alkane hydroxylase genes not currently annotated as alkane hydroxylase. Gene products show signs of optimization to low temperature, including regions of increased flexibility and amino acid preferences typical of psychrophilic proteins. These findings are consistent with observations of microbial degradation of crude oil in cold environments and identify proteins that can be targeted in rate studies and in the design of molecular tools for low temperature bioremediation.
Topics: Alkanes; Biocatalysis; Cold Temperature; Cytochrome P-450 CYP4A; Genomics; Phylogeny
PubMed: 25515036
DOI: 10.1186/1471-2164-15-1120 -
Zeitschrift Fur Naturforschung. C,... 2010An n-hexane extract of fresh, mature leaves of Argemone mexicana (Papaveraceae), containing thin-layer epicuticular waxes, has been analysed for the first time by TLC,...
An n-hexane extract of fresh, mature leaves of Argemone mexicana (Papaveraceae), containing thin-layer epicuticular waxes, has been analysed for the first time by TLC, IR and GLC using standard hydrocarbons. Seventeen long-chain alkanes (n-C18 to n-C34) were identified and quantified. Nonacosane (n-C29) was established as the n-alkane with the highest amount, whilst octadecane (n-C19) was the least abundant component of the extracted wax fraction. The carbon preference index (CPI) calculated for the hydrocarbon sample with the chain lengths between C18 and C34 was 1.2469, showing an odd to even carbon number predominance.
Topics: Alkanes; Argemone; India; Plant Extracts; Plant Leaves
PubMed: 21138052
DOI: 10.1515/znc-2010-9-1001 -
Nature Structural & Molecular Biology Apr 2023The cryo-EM structure of a natural AlkB–AlkG fusion from reveals the mechanistic basis for its selectivity towards, and functionalization of, alkane terminal C–H...
The cryo-EM structure of a natural AlkB–AlkG fusion from reveals the mechanistic basis for its selectivity towards, and functionalization of, alkane terminal C–H groups. AlkB contains an alkane entry tunnel and a diiron active site, and AlkG docks through electrostatic interactions and transfers electrons to the diiron center for catalysis.
Topics: Alkanes; Phylogeny
PubMed: 37002482
DOI: 10.1038/s41594-023-00964-2 -
Analytical Chemistry Aug 2021Power-to-gas is a heavily discussed option to store surplus electricity from renewable sources. Part of the generated hydrogen could be fed into the gas grid and lead to...
Power-to-gas is a heavily discussed option to store surplus electricity from renewable sources. Part of the generated hydrogen could be fed into the gas grid and lead to fluctuations in the composition of the fuel gas. Consequently, both operators of transmission networks and end users would need to frequently monitor the gas to ensure safety as well as optimal and stable operation. Currently, gas chromatography-based analysis methods are the state of the art. However, these methods have several downsides for time-resolved and distributed application and Raman gas spectroscopy is favorable for future point-of-use monitoring. Here, we demonstrate that fiber-enhanced Raman gas spectroscopy (FERS) enables the simultaneous detection of all relevant gases, from major (methane, CH; hydrogen, H) to minor (C2-C6 alkanes) fuel gas components. The characteristic peaks of H (585 cm), CH (2917 cm), isopentane (765 cm), -butane (798 cm), -butane (830 cm), -pentane (840 cm), propane (869 cm), ethane (993 cm), and -hexane (1038 cm) are well resolved in the broadband spectra acquired with a compact spectrometer. The fiber enhancement achieved in a hollow-core antiresonant fiber enables highly sensitive measurements with limits of detection between 90 and 180 ppm for different hydrocarbons. Both methane and hydrogen were quantified with high accuracy with average relative errors of 1.1% for CH and 1.5% for H over a wide concentration range. These results show that FERS is ideally suited for comprehensive fuel gas analysis in a future, where regenerative sources lead to fluctuations in the composition of gas.
Topics: Alkanes; Gases; Hydrogen; Methane; Spectrum Analysis, Raman
PubMed: 34297525
DOI: 10.1021/acs.analchem.1c01500 -
Biodegradation Apr 2022Application of genetically engineered bacterial strains for biodegradation of hydrocarbons is a sustainable solution for treating pollutants as well as in industrial...
Application of genetically engineered bacterial strains for biodegradation of hydrocarbons is a sustainable solution for treating pollutants as well as in industrial applications. However, the process of bioengineering should be carefully carried out to optimize the output. Investigation of regulatory genes for bioengineering is essential for developing synthetic circuits for effective biocatalysts. Here we focus on LcaR, a putative transcriptional regulator affecting the expression of alkB2 and lcaR operon that has a high potential to become a tool in designing such pathways. Four LcaR dimers bind specifically to the upstream regulatory region where divergent promoters of alkB2 and lcaR genes are located with high affinity at a K of 0.94 ± 0.17 nM and a Hill coefficient is 1.7 ± 0.3 demonstrating cooperativity in the association. Ligand binding alters the conformation of LcaR, which releases the regulator from its cognate DNA. Tetradecanal and hexadecanal act as natural ligands of LcaR with an IC values of 3.96 ± 0.59 µg/ml and 0.68 ± 0.21 µg/ml, respectively. The structure and function of transcription factors homologous to LcaR have not been characterized to date. This study provides insight into regulatory mechanisms of alkane degradation with a direction towards potential applications in bioengineering for bioremediation and industrial applications.
Topics: Alkanes; Bacterial Proteins; Biodegradation, Environmental; Bioengineering; Pseudomonas aeruginosa
PubMed: 34989928
DOI: 10.1007/s10532-021-09970-x -
Environmental Microbiology Dec 2022Alkane constitutes major fractions of crude oils, and its microbial aerobic degradation dominantly follows the terminal oxidation and the sub-terminal pathways. However,...
Alkane constitutes major fractions of crude oils, and its microbial aerobic degradation dominantly follows the terminal oxidation and the sub-terminal pathways. However, the latter one received much less attention, especially since the related genes were yet to be fully defined. Here, we isolated a bacterium designated Acinetobacter sp. strain NyZ410, capable of growing on alkanes with a range of chain lengths and derived sub-terminal oxidation products. From its genome, a secondary alcohol degradation gene cluster (sad) was identified to be likely involved in converting the aliphatic secondary alcohols (the sub-terminal oxidation products of alkanes) to the corresponding primary alcohols by removing two-carbon unit. On this cluster, sadC encoded an alcohol dehydrogenase converting the aliphatic secondary alcohols to the corresponding ketones; sadD encoded a Baeyer-Villiger monooxygenase catalysing the conversion of the aliphatic ketones to the corresponding esters; SadA and SadB are two esterases hydrolyzing aliphatic esters to the primary alcohols and acetic acids. Bioinformatics analyses indicated that the sad cluster was widely distributed in the genomes of probable alkane degraders, apparently coexisting (64%) with the signature enzymes AlkM and AlmA for alkane terminal oxidation in 350 bacterial genomes. It suggests that the alkane sub-terminal oxidation may be more ubiquitous than previously thought.
Topics: Alkanes; Alcohols; Ketones; Multigene Family; Esters
PubMed: 35837858
DOI: 10.1111/1462-2920.16124 -
Annual Review of Microbiology Sep 2022Alkanes are saturated apolar hydrocarbons that range from their simplest form, methane, to high-molecular-weight compounds. Although alkanes were once considered... (Review)
Review
Alkanes are saturated apolar hydrocarbons that range from their simplest form, methane, to high-molecular-weight compounds. Although alkanes were once considered biologically recalcitrant under anaerobic conditions, microbiological investigations have now identified several microbial taxa that can anaerobically degrade alkanes. Here we review recent discoveries in the anaerobic oxidation of alkanes with a specific focus on archaea that use specific methyl coenzyme M reductases to activate their substrates. Our understanding of the diversity of uncultured alkane-oxidizing archaea has expanded through the use of environmental metagenomics and enrichment cultures of syntrophic methane-, ethane-, propane-, and butane-oxidizing marine archaea with sulfate-reducing bacteria. A recently cultured group of archaea directly couples long-chain alkane degradation with methane formation, expanding the range of substrates used for methanogenesis. This article summarizes the rapidly growing knowledge of the diversity, physiology, and habitat distribution of alkane-degrading archaea.
Topics: Alkanes; Anaerobiosis; Archaea; Methane; Oxidation-Reduction; Phylogeny
PubMed: 35917471
DOI: 10.1146/annurev-micro-111021-045911 -
Microbial Cell Factories Nov 2020The development of sustainable routes to the bio-manufacture of gaseous hydrocarbons will contribute widely to future energy needs. Their realisation would contribute... (Review)
Review
The development of sustainable routes to the bio-manufacture of gaseous hydrocarbons will contribute widely to future energy needs. Their realisation would contribute towards minimising over-reliance on fossil fuels, improving air quality, reducing carbon footprints and enhancing overall energy security. Alkane gases (propane, butane and isobutane) are efficient and clean-burning fuels. They are established globally within the transportation industry and are used for domestic heating and cooking, non-greenhouse gas refrigerants and as aerosol propellants. As no natural biosynthetic routes to short chain alkanes have been discovered, de novo pathways have been engineered. These pathways incorporate one of two enzymes, either aldehyde deformylating oxygenase or fatty acid photodecarboxylase, to catalyse the final step that leads to gas formation. These new pathways are derived from established routes of fatty acid biosynthesis, reverse β-oxidation for butanol production, valine biosynthesis and amino acid degradation. Single-step production of alkane gases in vivo is also possible, where one recombinant biocatalyst can catalyse gas formation from exogenously supplied short-chain fatty acid precursors. This review explores current progress in bio-alkane gas production, and highlights the potential for implementation of scalable and sustainable commercial bioproduction hubs.
Topics: Alkanes; Biofuels; Biosynthetic Pathways; Butanes; Carboxy-Lyases; Fatty Acids; Gases; Genetic Engineering; Industrial Microbiology; Metabolic Engineering; Oxidation-Reduction; Oxygenases; Propane; Synthetic Biology
PubMed: 33187524
DOI: 10.1186/s12934-020-01470-6