Isolation and characterization of a gene encoding a S-adenosyl-l-methionine-dependent halide/thiol methyltransferase (HTMT) from the marine diatom Phaeodactylum tricornutum: Biogenic mechanism of CH(3)I emissions in oceans

Several marine algae including diatoms exhibit S-adenosyl-l-methionine (SAM) halide/thiol methyltransferase (HTMT) activity, which is involved in the emission of methyl halides. In this study, the in vivo biogenic emission of methyl iodide from the diatom Phaeodactylum tricornutum was found to be clearly correlated with iodide concentration in the incubation media. The gene encoding HTMT (Pthtmt) was isolated from P. tricornutum CCAP 1055/1, and expressed in Escherichia coli. The molecular weight of the enzyme was 29.7kDa including a histidine tag, and the optimal pH was around pH 7.0. The kinetic properties of recombinant PtHTMT towards Cl(-), Br(-), I(-), [SH](-), [SCN](-), and SAM were 637.88mM, 72.83mM, 8.60mM, 9.92mM, 7.9mM, and 0.016mM, respectively, and were similar to those of higher-plant HTMTs, except that the activity towards thiocyanate was lower. The biogenic emission of methyl halides from the cultured cells and the enzymatic properties of HTMT suggest that the HMT/HTMT reaction is key to understanding the biogenesis of methyl halides in oceanic environments as well as terrestrial ones.     

Biosynthesis of halomethanes and methanethiol by higher plants via a novel methyltransferase reaction

Biogenic emissions of halomethanes (CH₃CI, CH₃Br and CH₃I) and methanethiol (CH₃SH) are of major significance to atmospheric chemistry, but there is little information on such emissions from higher plants. We present evidence that plants can produce all these gases through an identical methyltransferase reaction. A survey of 118 herbaceous species, based on CH₃I production by leaf discs supplied with Kl, detected the presence of in vivo halide methyltransferase activity in 87 species. The activities ranged over nearly 4 orders of magnitude. Plants generally considered salt tolerant had relatively low activities, and salinization of three such species did not increase the activity. The highest activities were found in the family Brassicaceae. Leaf extracts of Brassica oleracea catalysed the S-adenosyl-L-methioninc-dependenl niethylalion of the halides I^?, Br^? and CI^? to the respective halomethanes. In addition, the extract similarly methylated HS^? (bisulphide) to CH₃SH. These two types of enzyme activity (halide and bisulphide methyltransferase) were also present in all of the 20 species comprising a subsample that represented the range of CH₃I emissions observed in the initial survey of in vivo CH₃I production ability, and in a marine red alga Endocladia muricata. Moreover, the two activities occurred in approximately the same ratio in all the higher plants tested. These findings highlight the potential of higher plants to contribute to the atmosphericbudget of halomethanes and melhanethiol. The halide and bisulphide methyltransferase activities may also provide a mechanism for the elimination of halide and HS^? ions, both of which are known to be phytotoxic.     

Environmental and biological controls on methyl halide emissions from southern California coastal salt marshes

Methyl bromide (CH₃Br) and methyl chloride(CH₃Cl) emission rates from southernCalifornia coastal salt marshes show largespatial and temporal variabilities that arestrongly linked to biological and environmentalfactors. Here we discuss biogeochemical linesof evidence pointing to vegetation as theprimary source of CH₃Br and CH₃Clemissions from salt marshes. Sediments andmacroalgae do not appear to be major producersof these compounds, based on observations thatthe highest fluxes are not inhibited by soilinundation; their emissions are not correlatedwith those of certain gases produced in soils;and emissions from mudflat- andmacroalgae-dominated sites are relativelysmall. In contrast, the seasonal and spatialvariabilities of methyl halide fluxes in thesesalt marshes are consistent with the productionof these compounds by vascular plants, althoughthe possibility of production by microflora orfungi associated with the salt marsh vegetationis not ruled out. Flux chamber measurements ofemission rates are largely correlated to theoverall plant biomass enclosed in the chamber,but appear also to be highly dependent on thepredominant plant species. Emission ratesfollow a diurnal trend similar to the trends ofambient air temperature and photosyntheticallyactive radiation, but not surface soiltemperature. Diurnal variabilities in thecarbon isotope compositions of CH₃Cl andCH₃Br and their relative ratios ofemissions are consistent with simultaneouslycompeting mechanisms of uptake andproduction.     

Synthesis and characterization of a series of N3O-ligated mononuclear Mn(II) halide complexes

Treatment of a N₃O-donor chelate ligand (mpppa = N-methyl-N-((6-pivaloylamido-2-pyridyl)methyl)-N-(2-pyridylethyl)amine; bpppa = N-benzyl-N-((6-pivaloylamido-2-pyridyl)methyl)-N-(2-pyridylmethyl)amine) with equimolar amounts of Mn(ClO₄)₂ · 6H₂O and Me₄NX (X = Cl, Br, I) in methanol resulted in the production of a series of mononuclear Mn(II) halide complexes of the formula [(L)Mn-X(CH₃OH)]ClO₄ (L = mpppa or bpppa). X-ray crystallographic studies of [(mpppa)Mn-Cl(CH₃OH)]ClO₄ · CH₃OH (2 · CH₃OH), [(mpppa)Mn-Br(CH₃OH)]ClO₄ · CH₃OH (4 · CH₃OH), [(mpppa)Mn-I(CH₃OH)]ClO₄ · CH₃OH (6 · CH₃OH), and [(bpppa)Mn-I(CH₃OH)]ClO₄ · O₂(CH₂CH₃)₂ (7 · O(CH₂CH₃)₂) revealed for each a mononuclear Mn(II) center having tetradentate coordination of the chelate ligand, one coordinated halide anion, and one molecule of coordinated methanol. An increase in the Mn-X distance through the halide series (Cl, Br, I) correlates linearly with the increase in the radius of the anion. The magnetic moment of each halide complex, measured via Evans method in methanol, is consistent with the presence of a high-spin distorted octahedral Mn(II) center. The EPR features of the halide complexes in methanol do not change as a function of the nature of the halide coordinated to the Mn(II) center.     

A review of bacterial methyl halide degradation: biochemistry,genetics and molecular ecology

Methyl halide-degrading bacteria are a diverse group of organisms that are found in both terrestrial and marine environments. They potentially play an important role in mitigating ozone depletion resulting from methyl chloride and methyl bromide emissions. The first step in the pathway(s) of methyl halide degradation involves a methyltransferase and, recently, the presence of this pathway has been studied in a number of bacteria. This paper reviews the biochemistry and genetics of methyl halide utilization in the aerobic bacteria Methylobacterium chloromethanicum CM4T, Hyphomicrobium chloromethanicum CM2T, Aminobacter strain IMB-1 and Aminobacter strain CC495. These bacteria are able to use methyl halides as a sole source of carbon and energy, are all members of the alpha-Proteobacteria and were isolated from a variety of polluted and pristine terrestrial environments. An understanding of the genetics of these bacteria identified a unique gene (cmuA) involved in the degradation of methyl halides, which codes for a protein (CmuA) with unique methyltransferase and corrinoid functions. This unique functional gene, cmuA, is being used to develop molecular ecology techniques to examine the diversity and distribution of methyl halide-utilizing bacteria in the environment and hopefully to understand their role in methyl halide degradation in different environments. These techniques will also enable the detection of potentially novel methyl halide-degrading bacteria.     

Theoretical studies on identity SN2 reactions of lithium halide and methyl halide: A microhydration model

Reactions of lithium halide (LiX, X = F, Cl, Br and I) and methyl halide (CH₃X, X = F, Cl, Br and I) have been investigated at the B3LYP/6-31G(d) level of theory using the microhydration model. Beginning with hydrated lithium ion, four or two water molecules have been conveniently introduced to these aqueous-phase halogen-exchange S_N2 reactions. These water molecules coordinated with the center metal lithium ion, and also interacted with entering and leaving halogen anion via hydrogen bond in complexes and transition state, which to some extent compensated hydration of halogen anion. At 298 K the reaction profiles all involve central barriers ΔE _cent which are found to decrease in the order F > Cl > Br > I. The same trend is also found for the overall barriers (ΔE _ovr ) of the title reaction. In the S_N2 reaction of sodium iodide and methyl iodide, the activation energy agrees well with the aqueous conductometric investigation.     

Fluorescence-Based Bacterial Bioreporter for Specific Detection of Methyl Halide Emissions in the Environment

Methyl halides are volatile one-carbon compounds responsible for substantial depletion of stratospheric ozone. Among them, chloromethane (CH₃Cl) is the most abundant halogenated hydrocarbon in the atmosphere. Global budgets of methyl halides in the environment are still poorly understood due to uncertainties in their natural sources, mainly from vegetation, and their sinks, which include chloromethane-degrading bacteria. A bacterial bioreporter for the detection of methyl halides was developed on the basis of detailed knowledge of the physiology and genetics of Methylobacterium extorquens CM4, an aerobic alphaproteobacterium which utilizes chloromethane as the sole source of carbon and energy. A plasmid construct with the promoter region of the chloromethane dehalogenase gene cmuA fused to a promotorless yellow fluorescent protein gene cassette resulted in specific methyl halide-dependent fluorescence when introduced into M. extorquens CM4. The bacterial whole-cell bioreporter allowed detection of methyl halides at femtomolar levels and quantification at concentrations above 10 pM (approximately 240 ppt). As shown for the model chloromethane-producing plant Arabidopsis thaliana in particular, the bioreporter may provide an attractive alternative to analytical chemical methods to screen for natural sources of methyl halide emissions.     

Analysis of genes involved in methyl halide degradation in Aminobacter lissarensis CC495

Aminobacter lissarensis CC495 is an aerobic facultative methylotroph capable of growth on glucose, glycerol, pyruvate and methylamine as well as the methyl halides methyl chloride and methyl bromide. Previously, cells grown on methyl chloride have been shown to express two polypeptides with apparent molecular masses of 67 and 29 kDa. The 67 kDa protein was purified and identified as a halomethane:bisulfide/halide ion methyltransferase. This study describes a single gene cluster in A. lissarensis CC495 containing the methyl halide utilisation genes cmuB, cmuA, cmuC, orf 188, paaE and hutI. The genes correspond to the same order and have a high similarity to a gene cluster found in Aminobacter ciceronei IMB-1 and Hyphomicrobium chloromethanicum strain CM2 indicating that genes encoding methyl halide degradation are highly conserved in these strains.     

Halomethane:Bisulfide/Halide Ion Methyltransferase, an Unusual Corrinoid Enzyme of Environmental Significance Isolated from an Aerobic Methylotroph Using Chloromethane as the Sole Carbon Source

A novel dehalogenating/transhalogenating enzyme, halomethane:bisulfide/halide ion methyltransferase, has been isolated from the facultatively methylotrophic bacterium strain CC495, which uses chloromethane (CH₃Cl) as the sole carbon source. Purification of the enzyme to homogeneity was achieved in high yield by anion-exchange chromatography and gel filtration. The methyltransferase was composed of a 67-kDa protein with a corrinoid-bound cobalt atom. The purified enzyme was inactive but was activated by preincubation with 5 mM dithiothreitol and 0.5 mM CH₃Cl; then it catalyzed methyl transfer from CH₃Cl, CH₃Br, or CH₃I to the following acceptor ions (in order of decreasing efficacy): I⁻, HS⁻, Cl⁻, Br⁻, NO₂⁻, CN⁻, and SCN⁻. Spectral analysis indicated that cobalt in the native enzyme existed as cob(II)alamin, which upon activation was reduced to the cob(I)alamin state and then was oxidized to methyl cob(III)alamin. During catalysis, the enzyme shuttles between the methyl cob(III)alamin and cob(I)alamin states, being alternately demethylated by the acceptor ion and remethylated by halomethane. Mechanistically the methyltransferase shows features in common with cobalamin-dependent methionine synthase from Escherichia coli. However, the failure of specific inhibitors of methionine synthase such as propyl iodide, N₂O, and Hg²⁺ to affect the methyltransferase suggests significant differences. During CH₃Cl degradation by strain CC495, the physiological acceptor ion for the enzyme is probably HS⁻, a hypothesis supported by the detection in cell extracts of methanethiol oxidase and formaldehyde dehydrogenase activities which provide a metabolic route to formate. 16S rRNA sequence analysis indicated that strain CC495 clusters with Rhizobium spp. in the alpha subdivision of the Proteobacteria and is closely related to strain IMB-1, a recently isolated CH₃Br-degrading bacterium (T. L. Connell Hancock, A. M. Costello, M. E. Lidstrom, and R. S. Oremland, Appl. Environ. Microbiol. 64:2899–2905, 1998). The presence of this methyltransferase in bacterial populations in soil and sediments, if widespread, has important environmental implications.     

Hydrologic regulation of gross methyl chloride and methyl bromide uptake from Alaskan Arctic tundra

The Arctic tundra has been shown to be a potentially significant regional sink for methyl chloride (CH₃Cl) and methyl bromide (CH₃Br), although prior field studies were spatially and temporally limited, and did not include gross flux measurements. Here we compare net and gross CH₃Cl and CH₃Br fluxes in the northern coastal plain and continental interior. As expected, both regions were net sinks for CH₃Cl and CH₃Br. Gross uptake rates (−793 nmol CH₃Cl m⁻² day⁻¹ and −20.3 nmol CH₃Br m⁻² day⁻¹) were 20–240% greater than net fluxes, suggesting that the Arctic is an even greater sink than previously believed. Hydrology was the principal regulator of methyl halide flux, with an overall trend towards increasing methyl halide uptake with decreasing soil moisture. Water table depth was one of the best predictors of net and gross uptake, with uptake increasing proportionately with water table depth. In drier areas, gross uptake was very high, averaging −1201 nmol CH₃Cl m⁻² day⁻¹ and −34.9 nmol CH₃Br m⁻² day⁻¹; in flooded areas, gross uptake was significantly lower, averaging −61 nmol CH₃Cl m⁻² day⁻¹ and −2.3 nmol CH₃Br m⁻² day⁻¹. Net and gross uptake was greater in the continental interior than in the northern coastal plain, presumably due to drier inland conditions. Within certain microtopographic features (low‐ and high‐centered polygons), uptake rates were positively correlated with soil temperature, indicating that temperature played a secondary role in methyl halide uptake. Incubations suggested that the inverse relationship between water content and methyl halide uptake was the result of mass transfer limitation in saturated soils, rather than because of reduced microbial activity under anaerobic conditions. These findings have potential regional significance, as the Arctic is expected to become warmer and drier due to anthropogenic climate forcing, potentially enhancing the Arctic sink for CH₃Cl and CH₃Br.     

Genetic control of methyl halide production in Arabidopsis

Methyl chloride (CH(3)Cl) and methyl bromide (CH(3)Br) are the primary carriers of natural chlorine and bromine, respectively, to the stratosphere, where they catalyze the destruction of ozone, whereas methyl iodide (CH(3)I) influences aerosol formation and ozone loss in the boundary layer. CH(3)Br is also an agricultural pesticide whose use is regulated by international agreement. Despite the economic and environmental importance of these methyl halides, their natural sources and biological production mechanisms are poorly understood. Besides CH(3)Br fumigation, important sources include oceans, biomass burning, tropical plants, salt marshes, and certain crops and fungi. Here, we demonstrate that the model plant Arabidopsis thaliana produces and emits methyl halides and that the enzyme primarily responsible for the production is encoded by the HARMLESS TO OZONE LAYER (HOL) gene. The encoded protein belongs to a group of methyltransferases capable of catalyzing the S-adenosyl-L-methionine (SAM)-dependent methylation of chloride (Cl(-)), bromide (Br(-)), and iodide (I(-)) to produce methyl halides. In mutant plants with the HOL gene disrupted, methyl halide production is largely eliminated. A phylogenetic analysis with the HOL gene suggests that the ability to produce methyl halides is widespread among vascular plants. This approach provides a genetic basis for understanding and predicting patterns of methyl halide production by plants.     

Theoretical study of the triangular bonding complex formed by carbon tetrabromide, a halide, and a solvent molecule in the gas phase

MP2(full)/aug-cc-pVDZ(-PP) computations predict that new triangular bonding complexes (where X^? is a halide and H–C refers to a protic solvent molecule) consist of one halogen bond and two hydrogen bonds in the gas phase. Carbon tetrabromide acts as the donor in the halogen bond, while it acts as an acceptor in the hydrogen bond. The halide (which commonly acts as an acceptor) can interact with both carbon tetrabromide and solvent molecule (CH₃CN, CH₂Cl₂, CHCl₃) to form a halogen bond and a hydrogen bond, respectively. The strength of the halogen bond obeys the order CBr₄???Cl^? > CBr₄???Br^? > CBr₄???I^?. For the hydrogen bonds formed between various halides and the same solvent molecule, the strength of the hydrogen bond obeys the order C-H???Cl^? > C-H???Br^? > C-H???I^?. For the hydrogen bonds formed between the same halide and various solvent molecules, the interaction strength is proportional to the acidity of the hydrogen in the solvent molecule. The diminutive effect is present between the hydrogen bonds and the halogen bond in chlorine and bromine triangular bonding complexes. Complexes containing iodide ion show weak cooperative effects.

Effects of feed intake on enteric methane emissions from sheep fed fresh white clover (Trifolium repens) and perennial ryegrass (Lolium perenne) forages

Published analyses of enteric methane (CH₄) emissions from sheep and cattle show an inverse relationship between feed intake and CH₄ yield (g CH₄/kg dry matter (DM) intake), which suggests opportunities for reducing CH₄ emissions from feed eaten and per unit of animal production. Most relationships between feed intake and CH₄ yield have been based on animals fed conserved feeds, especially silages and grains. Our research is a series of experiments with fresh white clover (Trifolium repens) and perennial ryegrass (Lolium perenne; ryegrass) forages fed to sheep at a range of feed intake levels. This study was comprised of four experiments where good quality freshly harvested white clover or ryegrass were fed to sheep over a three-fold range in DM intake, and CH₄ emissions were measured in respiration chambers for two consecutive days in each experiment. Measurements were made from 16 sheep in Experiment 1 (fed at 1.6 × metabolizable energy requirements for maintenance; ME_m), 28 sheep in Experiment 2 (at 0.8 and 2.0 × ME_m), eight sheep and two measurement periods in Experiment 3 (at 1.6 × ME_m), and 30 sheep in Experiment 4 (fed at 0.8, 1.2, 1.6, 2.0 and 2.5 × ME_m). Prior to each experiment, sheep had a 10 d acclimatization period to diets. Apparent digestibility was measured over 7 d from sheep in Experiments 1, 3 and 4, along with collection of rumen digesta for volatile fatty acid (VFA) determination. Although CH₄ yields differed when sheep were fed white clover or ryegrass at similar intakes, the differences were inconsistent and mean values similar across all experiments. This, and a similar structure of all experiments, enabled combined analysis of data from all four experiments using the restricted maximum likelihood (REML) procedure to estimate effects of feed intake level on digestibility, digestible nutrient intake, gas emissions, and VFA concentrations in the rumen. The REML analysis showed that when DM intake increased from 0.40 to 1.60 kg/d, the predicted responses were an increase in CH₄ production (g/d) of 187% (12.4–35.6 g/d; P<0.001), and a decline in CH₄ yield of 21% (25.6–20.2 g/kg DM intake; P<0.001). High feed intake levels were associated with increased molar proportions (mM of total VFA) of propionate from 0.17 to 0.21 (P=0.038). Single and multiple regressions were completed on the data from all experiments, with organic matter (OM) intake predicting 0.87 of the variation in CH₄ production, and molar proportion of propionate predicting 0.60 of the variation in CH₄ yield. Increasing feed intakes by 1 kg/d of DM reduced CH₄ yield by 4.5 g/kg DM intake. Plant chemical composition was weakly related to CH₄ yield. High intakes of fresh forages will lower CH₄ yield from fermentation, but effects of feed composition on CH₄ emissions were minor. The interaction between effects of feed intake and rumen function requires further investigation to understand relationships with CH₄ emissions.     

Inundation strongly stimulates nitrous oxide emissions from stems of the upland tree Fagus sylvatica and the riparian tree Alnus glutinosa

Background and aims

Nitrous oxide (N₂O) and methane (CH₄) can be emitted from surfaces of riparian plants. Data on the emission of these greenhouse gases by upland trees are scarce. We quantified CH₄ and N₂O emissions from stems of Fagus sylvatica, an upland tree, and Alnus glutinosa, a riparian tree.

Methods

The gas fluxes were investigated in mesocosms under non-flooded control conditions and during a flooding period using static chamber systems and gas chromatographic analyses.

Results

Despite differences in the presence of an aerenchyma system, both tree species emitted N₂O and CH₄ from the stems. Flooding caused a dramatic transient increase of N₂O stem emissions by factors of 740 (A. glutinosa) and even 14,230 (F. sylvatica). Stem emissions of CH₄ were low and even deposition was determined (F. sylvatica controls). The results suggest that CH₄ was transported mainly through the aerenchyma, whereas N₂O transport occurred in the xylem sap.

Conclusions

For the first time it has been demonstrated that upland trees such as F. sylvatica clearly significantly emit N₂O from their stems despite lacking an aerenchyma. If this result is confirmed in adult trees, upland forests may constitute a new and significant source of atmospheric N₂O.     

Arabidopsis HARMLESS TO OZONE LAYER Protein Methylates a Glucosinolate Breakdown Product and Functions in Resistance to Pseudomonas syringae pv. maculicola

Almost all of the chlorine-containing gas emitted from natural sources is methyl chloride (CH₃Cl), which contributes to the destruction of the stratospheric ozone layer. Tropical and subtropical plants emit substantial amounts of CH₃Cl. A gene involved in CH₃Cl emission from Arabidopsis was previously identified and designated HARMLESS TO OZONE LAYER (hereafter AtHOL1) based on the mutant phenotype. Our previous studies demonstrated that AtHOL1 and its homologs, AtHOL2 and AtHOL3, have S-adenosyl-l-methionine-dependent methyltransferase activities. However, the physiological functions of AtHOLs have yet to be elucidated. In the present study, our comparative kinetic analyses with possible physiological substrates indicated that all of the AtHOLs have low activities toward chloride. AtHOL1 was highly reactive to thiocyanate (NCS⁻), a pseudohalide, synthesizing methylthiocyanate (CH₃SCN) with a very high k_cat/K_m value. We demonstrated in vivo that substantial amounts of NCS⁻ were synthesized upon tissue damage in Arabidopsis and that NCS⁻ was largely derived from myrosinase-mediated hydrolysis of glucosinolates. Analyses with the T-DNA insertion Arabidopsis mutants (hol1, hol2, and hol3) revealed that only hol1 showed increased sensitivity to NCS⁻ in medium and a concomitant lack of CH₃SCN synthesis upon tissue damage. Bacterial growth assays indicated that the conversion of NCS⁻ into CH₃SCN dramatically increased antibacterial activities against Arabidopsis pathogens that normally invade the wound site. Furthermore, hol1 seedlings showed an increased susceptibility toward an Arabidopsis pathogen, Pseudomonas syringae pv. maculicola. Here we propose that AtHOL1 is involved in glucosinolate metabolism and defense against phytopathogens. Moreover, CH₃Cl synthesized by AtHOL1 could be considered a byproduct of NCS⁻ metabolism.Methyl chloride (CH₃Cl) is the most abundant halohydrocarbon emitted into the atmosphere and constitutes about 17% of the chlorine currently in the stratosphere (1). CH₃Cl is derived mainly from natural sources and contributes to the destruction of the stratospheric ozone layer. As the total abundance of ozone-depleting gases such as chlorofluorocarbons in the atmosphere has begun to decrease in recent years as a result of The Montreal Protocol on Substances That Deplete the Ozone Layer, the impact of CH₃Cl emission from natural sources will become greater on the atmospheric chemistry. CH₃Cl emission into the atmosphere has been estimated at 1,700–13,600 Gg/year (1), which underscores the great uncertainty of the estimation. Oceans (2), biomass burning (3), wood-rotting fungi, and coastal salt marshes (4) are the major sources of CH₃Cl production. Recently, it was reported that large amounts of CH₃Cl are emitted from tropical and subtropical plants, which are hence considered as the major sources of CH₃Cl (5–7). It was estimated that the CH₃Cl emission from tropical plants could account for 30–50% of the global CH₃Cl emission (8). To accomplish an accurate estimation of CH₃Cl production in the atmosphere through “bottom-up” approaches, elucidating the mechanisms and physiological functions of CH₃Cl emission from plants will be important.The biological synthesis of methyl halides has been demonstrated mainly by biochemical analyses. The enzymatic activities that transfer a methyl group from S-adenosyl-l-methionine (SAM)² to halide ions (Cl⁻, Br⁻, I⁻), which synthesize methyl halides, were first discovered in cell-free extracts of Phellinus pomaceus (a white rot fungus), Endocladia muricata (a marine red alga), and Mesembryanthemum crystallinum (ice plant, a halophytic plant) (9). Enzyme purification and cDNA cloning of the methyl chloride transferase (MCT) was first reported with Batis maritima, a halophytic plant that grows abundantly in salt marshes. As high concentrations of ions such as Cl⁻ are often detrimental to plants, halophytic plants are considered to possess various salt tolerance mechanisms. MCT was hypothesized to control and regulate the internal concentration of Cl⁻, rich in the habitat in which halophytic plant grows (10, 11).In the meantime, purification of thiol methyltransferase (TMT), which methylates bisulfide (HS⁻) and halide (Cl⁻, Br⁻, I⁻) ions was reported with cabbage, Brassica oleracea (12). The purified and recombinant TMTs were later shown to also methylate the thiocyanate ion (NCS⁻), which is called pseudohalide because of its chemical properties similar to halide ions (13, 14). NCS⁻ is a hydrolysis product found in some glucosinolates, which are secondary metabolites found mainly in the order Brassicales including the model plant Arabidopsis thaliana (15). Upon tissue damage such as by insect or herbivore attack, glucosinolates are hydrolyzed by myrosinase (β-thioglucosidase) into biologically active compounds including isothiocyanates. Isothiocyanates derived from indole glucosinolates and 4-hydroxybenzyl glucosinolates are reported to be highly unstable and yield NCS⁻ upon reacting with various nucleophiles (15–17). Based on the enzymatic activity, the physiological role of TMT was speculated to metabolize glucosinolate breakdown products (14). However, there are no reported studies that examine these MCT and TMT hypotheses through in vivo experiments.An Arabidopsis homolog of MCT was also identified, and its T-DNA insertion Arabidopsis mutants were analyzed (18). Because the gene disruption eliminated almost all of the methyl halide emissions from the mutants, the gene was revealed to be involved in methyl halide synthesis and was designated HOL (HARMLESS TO OZONE LAYER; denoted as AtHOL1 in our studies) based on the mutant phenotype (18). Recently, we identified AtHOL1 homologs AtHOL2 and AtHOL3 in Arabidopsis, and we demonstrated biochemically that the three recombinant AtHOLs have SAM-dependent methyltransferase activities (19). In this study, reverse genetic and biochemical analyses of all AtHOL isoforms revealed that AtHOL1 in vivo is involved in the methylation of NCS⁻ produced by glucosinolate hydrolysis. Although there are several studies that have examined the biological activities of glucosinolate hydrolysis products, the mechanisms of NCS⁻ synthesis and its methylation to methyl thiocyanate (CH₃SCN) have yet to be reported in detail. The biological activity and physiological function of CH₃SCN synthesized by AtHOL1 was also examined.     

Structural basis and mechanism of the inhibition of the type-3 copper protein tyrosinase from Streptomyces antibioticus by halide ions

The inhibition of the type-3 copper enzyme tyrosinase by halide ions was studied by kinetic and paramagnetic (1)H NMR methods. All halides are inhibitors in the conversion of l-3,4-dihydroxyphenylalanine (l-DOPA) with apparent inhibition constants that follow the order I(-) < F(-) < Cl(-) < Br(-) at pH 6.80. The results show that the inhibition arises from the interaction of halide with both the oxidized (affinity F(-) > Cl(-) > Br(-) > I(-)) and reduced (affinity I(-) > Br(-) > Cl(-) > F(-)) enzyme. The paramagnetic (1)H NMR of the oxidized enzyme complexed with the halides is consistent with a direct interaction of halide with the type-3 site and shows that the (Cu-His(3))(2) coordination occurs in all halide-bound species. It is surmised that halides bridge both of the copper ions in the active site. Fluoride and chloride are shown to bind only to the low pH form of oxidized tyrosinase, explaining the strong pH dependence of the inhibition by these ions. We further show that p-toluic acid and the bidentate transition state analogue, Kojic acid, displace chloride from the oxidized active site, whereas the monodentate substrate analogue, p-nitrophenol, forms a ternary complex with the enzyme and the chloride ion. On the basis of the experimental results, a model is formulated for the inhibitor action and for the reaction of diphenols with the oxidized enzyme.     

Characterizing spatiotemporal dynamics of methane emissions from rice paddies in Northeast China from 1990 to 2010

Background

Rice paddies have been identified as major methane (CH₄) source induced by human activities. As a major rice production region in Northern China, the rice paddies in the Three-Rivers Plain (TRP) have experienced large changes in spatial distribution over the recent 20 years (from 1990 to 2010). Consequently, accurate estimation and characterization of spatiotemporal patterns of CH₄ emissions from rice paddies has become an pressing issue for assessing the environmental impacts of agroecosystems, and further making GHG mitigation strategies at regional or global levels.

Methodology/Principal Findings

Integrating remote sensing mapping with a process-based biogeochemistry model, Denitrification and Decomposition (DNDC), was utilized to quantify the regional CH₄ emissions from the entire rice paddies in study region. Based on site validation and sensitivity tests, geographic information system (GIS) databases with the spatially differentiated input information were constructed to drive DNDC upscaling for its regional simulations. Results showed that (1) The large change in total methane emission that occurred in 2000 and 2010 compared to 1990 is distributed to the explosive growth in amounts of rice planted; (2) the spatial variations in CH₄ fluxes in this study are mainly attributed to the most sensitive factor soil properties, i.e., soil clay fraction and soil organic carbon (SOC) content, and (3) the warming climate could enhance CH₄ emission in the cool paddies.

Conclusions/Significance

The study concluded that the introduction of remote sensing analysis into the DNDC upscaling has a great capability in timely quantifying the methane emissions from cool paddies with fast land use and cover changes. And also, it confirmed that the northern wetland agroecosystems made great contributions to global greenhouse gas inventory.     

Seasonal variations of methane fluxes from an unvegetated tidal freshwater mudflat (Hammersmith Creek, GA)

Wetlands are estimated to contribute nearly 40 % of global annual methane (CH₄) emissions to the atmosphere. However, because CH₄ fluxes from these systems vary spatially, seasonally, and by wetland type, there is a large uncertainty associated with scaling up the CH₄ flux from these environments. We monitored seasonal patterns of CH₄ cycling from tidal mudflat wetland sediments adjacent to a vegetated freshwater wetland in coastal Georgia between 2008 and 2009. CH₄ emissions were significantly correlated with CH₄ production and sediment saturation state with respect to CH₄ but not with temperature. CH₄ cycling displayed distinct seasonal patterns. Winter months were characterized by low CH₄ production and emissions. During the spring, summer and fall, CH₄ fluxes exceeded CH₄ production in the top 40 cm. Comparison of CH₄ sources and sinks in conjunction with the interpretation of CH₄ concentration profiles using a 1D reactive transport model indicated that CH₄ delivered via lateral tidal pumping likely provided additional CH₄ to the upper sediment column. Seasonally high CH₄ ebullition rates reflected increased CH₄ production and decreased CH₄ solubility. The annual CH₄ flux was estimated to be on the order of 10 mol CH₄ m^?2 y^?1 which is 2–4 times the global average for wetland CH₄ emissions. Thus, even though tidal freshwater mudflats are of limited spatial extent, these environments may serve as globally significant sources of CH₄ to the atmosphere. This study highlights the importance of these dynamic environments to the global CH₄ cycle and their relevance to climate change.     

Tuning optical properties and optical rotation of 3‐mercaptopropionic acid capped organic–inorganic hybrid perovskites

The emission wavelength of organic–inorganic hybrid perovskite quantum dots (QDs) can be tuned by controlling reaction time relevant to the halide exchange. It is because halide exchange with different time would lead to different molar ratio of halides in perovskite QDs such as Cl and Br. Here, to research the ligand's effect on the halide exchange, this work synthesized 3‐mercaptopropionic acid (MPA)‐capped CH₃NH₃PbBr_xCl_3‐x QDs. It was found that SH^? of MPA appeared to inhibit the halide exchange during the reation. Moreover, although the MPA‐capped CH₃NH₃PbBr_xCl_3‐x QDs did not contain the chiral centre, they exhibit the optical rotation. This may provide a method for chirality manipulation of perovskite.