AMP-dependent phosphite dehydrogenase, a phosphorylating enzyme in dissimilatory phosphite oxidation.

Oxidation of phosphite (HPO32-) to phosphate (HPO42-) releases electrons at a very low redox potential (E0'= -690 mV) which renders phosphite an excellent electron donor for microbial energy metabolism. To date, two pure cultures of strictly anaerobic bacteria have been isolated that run their energy metabolism on the basis of phosphite oxidation, the Gram-negative Desulfotignum phosphitoxidans (DSM 13687) and the Gram-positive Phosphitispora fastidiosa (DSM 112739). Here, we describe the key enzyme for dissimilatory phosphite oxidation in these bacteria. The enzyme catalyzed phosphite oxidation in the presence of adenosine monophosphate (AMP) to form adenosine diphosphate (ADP), with concomitant reduction of oxidized nicotinamide adenine dinucleotide (NAD+) to reduced nicotinamide adenine dinucleotide (NADH). The enzyme of P. fastidiosa was heterologously expressed in Escherichia coli. It has a molecular mass of 35.2 kDa and a high affinity for phosphite and NAD+. Its activity was enhanced more than 100-fold by addition of ADP-consuming adenylate kinase (myokinase) to a maximal activity between 30 and 80 mU x mg protein-1. A similar NAD-dependent enzyme oxidizing phosphite to phosphate with concomitant phosphorylation of AMP to ADP is found in D. phosphitoxidans, but this enzyme could not be heterologously expressed. Based on sequence analysis, these phosphite-oxidizing enzymes are related to nucleotide-diphosphate-sugar epimerases and indeed represent AMP-dependent phosphite dehydrogenases (ApdA). A reaction mechanism is proposed for this unusual type of substrate-level phosphorylation reaction.

Anaerobic dissimilatory phosphite oxidation, an extremely efficient concept of microbial electron economy.

Phosphite is a stable phosphorus compound that, together with phosphate, made up a substantial part of the total phosphorus content of the prebiotic Earth's crust. Oxidation of phosphite to phosphate releases electrons at an unusually low redox potential (-690 mV at pH 7.0). Numerous aerobic and anaerobic bacteria use phosphite as a phosphorus source and oxidise it to phosphate for synthesis of nucleotides and other phosphorus-containing cell constituents. Only two pure cultures of strictly anaerobic bacteria have been isolated so far that use phosphite as an electron donor in their energy metabolism, the Gram-positive Phosphitispora fastidiosa and the Gram-negative Desulfotignum phosphitoxidans. The key enzyme of this metabolism is an NAD+ -dependent phosphite dehydrogenase enzyme that phosphorylates AMP to ADP. These phosphorylating phosphite dehydrogenases were found to be related to nucleoside diphosphate sugar epimerases. The produced NADH is channelled into autotrophic CO2 fixation via the Wood-Ljungdahl (CO-DH) pathway, thus allowing for nearly complete assimilation of the substrate electrons into bacterial biomass. This extremely efficient type of electron flow connects energy and carbon metabolism directly through NADH and might have been important in the early evolution of life when phosphite was easily available on Earth.

Biodegradable High-Density Polyethylene-like Material.

We report a novel polyester material generated from readily available biobased 1,18-octadecanedicarboxylic acid and ethylene glycol possesses a polyethylene-like solid-state structure and also tensile properties similar to high density polyethylene (HDPE). Despite its crystallinity, high melting point (Tm =96 °C) and hydrophobic nature, polyester-2,18 is subject to rapid and complete hydrolytic degradation in in vitro assays with isolated naturally occurring enzymes. Under industrial composting conditions (ISO standard 14855-1) the material is biodegraded with mineralization above 95 % within two months. Reference studies with polyester-18,18 (Tm =99 °C) reveal a strong impact of the nature of the diol repeating unit on degradation rates, possibly related to the density of ester groups in the amorphous phase. Depolymerization by methanolysis indicates suitability for closed-loop recycling.

A glycyl radical enzyme enables hydrogen sulfide production by the human intestinal bacterium Bilophila wadsworthia.

Hydrogen sulfide (H2S) production in the intestinal microbiota has many contributions to human health and disease. An important source of H2S in the human gut is anaerobic respiration of sulfite released from the abundant dietary and host-derived organic sulfonate substrate in the gut, taurine (2-aminoethanesulfonate). However, the enzymes that allow intestinal bacteria to access sulfite from taurine have not yet been identified. Here we decipher the complete taurine desulfonation pathway in Bilophila wadsworthia 3.1.6 using differential proteomics, in vitro reconstruction with heterologously produced enzymes, and identification of critical intermediates. An initial deamination of taurine to sulfoacetaldehyde by a known taurine:pyruvate aminotransferase is followed, unexpectedly, by reduction of sulfoacetaldehyde to isethionate (2-hydroxyethanesulfonate) by an NADH-dependent reductase. Isethionate is then cleaved to sulfite and acetaldehyde by a previously uncharacterized glycyl radical enzyme (GRE), isethionate sulfite-lyase (IslA). The acetaldehyde produced is oxidized to acetyl-CoA by a dehydrogenase, and the sulfite is reduced to H2S by dissimilatory sulfite reductase. This unique GRE is also found in Desulfovibrio desulfuricans DSM642 and Desulfovibrio alaskensis G20, which use isethionate but not taurine; corresponding knockout mutants of D. alaskensis G20 did not grow with isethionate as the terminal electron acceptor. In conclusion, the novel radical-based C-S bond-cleavage reaction catalyzed by IslA diversifies the known repertoire of GRE superfamily enzymes and enables the energy metabolism of B. wadsworthia This GRE is widely distributed in gut bacterial genomes and may represent a novel target for control of intestinal H2S production.

Activation of short-chain ketones and isopropanol in sulfate-reducing bacteria.

BACKGROUND: Degradation of acetone by aerobic and nitrate-reducing bacteria can proceed via carboxylation to acetoacetate and subsequent thiolytic cleavage to two acetyl residues. A different strategy was identified in the sulfate-reducing bacterium Desulfococcus biacutus that involves formylation of acetone to 2-hydroxyisobutyryl-CoA. RESULTS: Utilization of short-chain ketones (acetone, butanone, 2-pentanone and 3-pentanone) and isopropanol by the sulfate reducer Desulfosarcina cetonica was investigated by differential proteome analyses and enzyme assays. Two-dimensional protein gel electrophoresis indicated that D. cetonica during growth with acetone expresses enzymes homologous to those described for Desulfococcus biacutus: a thiamine diphosphate (TDP)-requiring enzyme, two subunits of a B12-dependent mutase, and a NAD+-dependent dehydrogenase. Total proteomics of cell-free extracts confirmed these results and identified several additional ketone-inducible proteins. Acetone is activated, most likely mediated by the TDP-dependent enzyme, to a branched-chain CoA-ester, 2-hydroxyisobutyryl-CoA. This compound is linearized to 3-hydroxybutyryl-CoA by a coenzyme B12-dependent mutase followed by oxidation to acetoacetyl-CoA by a dehydrogenase. Proteomic analysis of isopropanol- and butanone-grown cells revealed the expression of a set of enzymes identical to that expressed during growth with acetone. Enzyme assays with cell-free extract of isopropanol- and butanone-grown cells support a B12-dependent isomerization. After growth with 2-pentanone or 3-pentanone, similar protein patterns were observed in cell-free extracts as those found after growth with acetone. CONCLUSIONS: According to these results, butanone and isopropanol, as well as the two pentanone isomers, are degraded by the same enzymes that are used also in acetone degradation. Our results indicate that the degradation of several short-chain ketones appears to be initiated by TDP-dependent formylation in sulfate-reducing bacteria.

Bacterial microcompartments for isethionate desulfonation in the taurine-degrading human-gut bacterium Bilophila wadsworthia.

BACKGROUND: Bilophila wadsworthia, a strictly anaerobic, sulfite-reducing bacterium and common member of the human gut microbiota, has been associated with diseases such as appendicitis and colitis. It is specialized on organosulfonate respiration for energy conservation, i.e., utilization of dietary and host-derived organosulfonates, such as taurine (2-aminoethansulfonate), as sulfite donors for sulfite respiration, producing hydrogen sulfide (H2S), an important intestinal metabolite that may have beneficial as well as detrimental effects on the colonic environment. Its taurine desulfonation pathway involves the glycyl radical enzyme (GRE) isethionate sulfite-lyase (IslAB), which cleaves isethionate (2-hydroxyethanesulfonate) into acetaldehyde and sulfite. RESULTS: We demonstrate that taurine metabolism in B. wadsworthia 3.1.6 involves bacterial microcompartments (BMCs). First, we confirmed taurine-inducible production of BMCs by proteomic, transcriptomic and ultra-thin sectioning and electron-microscopical analyses. Then, we isolated BMCs from taurine-grown cells by density-gradient ultracentrifugation and analyzed their composition by proteomics as well as by enzyme assays, which suggested that the GRE IslAB and acetaldehyde dehydrogenase are located inside of the BMCs. Finally, we are discussing the recycling of cofactors in the IslAB-BMCs and a potential shuttling of electrons across the BMC shell by a potential iron-sulfur (FeS) cluster-containing shell protein identified by sequence analysis. CONCLUSIONS: We characterized a novel subclass of BMCs and broadened the spectrum of reactions known to take place enclosed in BMCs, which is of biotechnological interest. We also provided more details on the energy metabolism of the opportunistic pathobiont B. wadsworthia and on microbial H2S production in the human gut.

Anthropogenic impact on the historical phytoplankton community of Lake Constance reconstructed by multimarker analysis of sediment-core environmental DNA.

During the 20th century, many lakes in the Northern Hemisphere were affected by increasing human population and urbanization along their shorelines and catchment, resulting in aquatic eutrophication. Ecosystem monitoring commenced only after the changes became apparent, precluding any examination of timing and dynamics of initial community change in the past and comparison of pre- and postimpact communities. Peri-Alpine Lake Constance (Germany) underwent a mid-century period of eutrophication followed by re-oligotrophication since the 1980s and is now experiencing warm temperatures. We extended the period for which monitoring data of indicator organisms exist by analysing historical environmental DNA (eDNA) from a sediment core dating back some 110 years. Using three metabarcoding markers-for microbial eukaryotes, diatoms and cyanobacteria-we revealed two major breakpoints of community change, in the 1930s and the mid-1990s. In our core, the latest response was exhibited by diatoms, which are classically used as palaeo-bioindicators for the trophic state of lakes. Following re-oligotrophication, overall diversity values reverted to similar ones of the early 20th century, but multivariate analysis indicated that the present community is substantially dissimilar. Community changes of all three groups were strongly correlated to phosphorus concentration changes, whereas significant relationships to temperature were only observed when we did not account for temporal autocorrelation. Our results indicate that each microbial group analysed exhibited a unique response, highlighting the particular strength of multimarker analysis of eDNA, which is not limited to organisms with visible remains and can therefore discover yet unknown responses and abiotic-biotic relationships.

Phosphitispora fastidiosa gen. nov. sp. nov., a new dissimilatory phosphite-oxidizing anaerobic bacterium isolated from anaerobic sewage sludge.

A new strictly anaerobic bacterium, strain DYL19T, was enriched and isolated with phosphite as the sole electron donor and CO2 as a single carbon source and electron acceptor from anaerobic sewage sludge sampled at a sewage treatment plant in Constance, Germany. It is a Gram-positive, spore-forming, slightly curved, rod-shaped bacterium which oxidizes phosphite to phosphate while reducing CO2 to biomass and small amounts of acetate. Optimal growth is observed at 30 °C, pH 7.2, with a doubling time of 3 days. Beyond phosphite, no further inorganic or organic electron donor can be used, and no other electron acceptor than CO2 is reduced. Sulphate inhibits growth with phosphite and CO2. The G+C content is 45.95 mol%, and dimethylmenaquinone-7 is the only quinone detectable in the cells. On the basis of 16S rRNA gene sequence analysis and other chemotaxonomic properties, strain DYL19T is described as the type strain of a new genus and species, Phosphitispora fastidiosa gen. nov., sp. nov.

Sulphoglycolysis in Escherichia coli K-12 closes a gap in the biogeochemical sulphur cycle.

Sulphoquinovose (SQ, 6-deoxy-6-sulphoglucose) has been known for 50 years as the polar headgroup of the plant sulpholipid in the photosynthetic membranes of all higher plants, mosses, ferns, algae and most photosynthetic bacteria. It is also found in some non-photosynthetic bacteria, and SQ is part of the surface layer of some Archaea. The estimated annual production of SQ is 10,000,000,000 tonnes (10 petagrams), thus it comprises a major portion of the organo-sulphur in nature, where SQ is degraded by bacteria. However, despite evidence for at least three different degradative pathways in bacteria, no enzymic reaction or gene in any pathway has been defined, although a sulphoglycolytic pathway has been proposed. Here we show that Escherichia coli K-12, the most widely studied prokaryotic model organism, performs sulphoglycolysis, in addition to standard glycolysis. SQ is catabolised through four newly discovered reactions that we established using purified, heterologously expressed enzymes: SQ isomerase, 6-deoxy-6-sulphofructose (SF) kinase, 6-deoxy-6-sulphofructose-1-phosphate (SFP) aldolase, and 3-sulpholactaldehyde (SLA) reductase. The enzymes are encoded in a ten-gene cluster, which probably also encodes regulation, transport and degradation of the whole sulpholipid; the gene cluster is present in almost all (>91%) available E. coli genomes, and is widespread in Enterobacteriaceae. The pathway yields dihydroxyacetone phosphate (DHAP), which powers energy conservation and growth of E. coli, and the sulphonate product 2,3-dihydroxypropane-1-sulphonate (DHPS), which is excreted. DHPS is mineralized by other bacteria, thus closing the sulphur cycle within a bacterial community.

Regulatory role of CsqR (YihW) in transcription of the genes for catabolism of the anionic sugar sulfoquinovose (SQ) in Escherichia coli K-12.

The binding sites of YihW, an uncharacterized DeoR-family transcription factor (TF) of Escherichia coli K-12, were identified using Genomic SELEX screening at two closely located sites, one inside the spacer between the bidirectional transcription units comprising the yihUTS operon and the yihV gene, and another one upstream of the yihW gene itself. Recently the YihUTS and YihV proteins were identified as catalysing the catabolism of sulfoquinovose (SQ), a hydrolysis product of sulfoquinovosyl diacylglycerol (SQDG) derived from plants and other photosynthetic organisms. Gel shift assay in vitro and reporter assay in vivo indicated that YihW functions as a repressor for all three transcription units. De-repression of the yih operons was found to be under the control of SQ as inducer, but not of lactose, glucose or galactose. Furthermore, a mode of its cooperative DNA binding was suggested for YihW by atomic force microscopy. Hence, as a regulator of the catabolism of SQ, we renamed YihW as CsqR.

Entner-Doudoroff pathway for sulfoquinovose degradation in Pseudomonas putida SQ1.

Sulfoquinovose (SQ; 6-deoxy-6-sulfoglucose) is the polar head group of the plant sulfolipid SQ-diacylglycerol, and SQ comprises a major proportion of the organosulfur in nature, where it is degraded by bacteria. A first degradation pathway for SQ has been demonstrated recently, a "sulfoglycolytic" pathway, in addition to the classical glycolytic (Embden-Meyerhof) pathway in Escherichia coli K-12; half of the carbon of SQ is abstracted as dihydroxyacetonephosphate (DHAP) and used for growth, whereas a C3-organosulfonate, 2,3-dihydroxypropane sulfonate (DHPS), is excreted. The environmental isolate Pseudomonas putida SQ1 is also able to use SQ for growth, and excretes a different C3-organosulfonate, 3-sulfolactate (SL). In this study, we revealed the catabolic pathway for SQ in P. putida SQ1 through differential proteomics and transcriptional analyses, by in vitro reconstitution of the complete pathway by five heterologously produced enzymes, and by identification of all four organosulfonate intermediates. The pathway follows a reaction sequence analogous to the Entner-Doudoroff pathway for glucose-6-phosphate: It involves an NAD(+)-dependent SQ dehydrogenase, 6-deoxy-6-sulfogluconolactone (SGL) lactonase, 6-deoxy-6-sulfogluconate (SG) dehydratase, and 2-keto-3,6-dideoxy-6-sulfogluconate (KDSG) aldolase. The aldolase reaction yields pyruvate, which supports growth of P. putida, and 3-sulfolactaldehyde (SLA), which is oxidized to SL by an NAD(P)(+)-dependent SLA dehydrogenase. All five enzymes are encoded in a single gene cluster that includes, for example, genes for transport and regulation. Homologous gene clusters were found in genomes of other P. putida strains, in other gamma-Proteobacteria, and in beta- and alpha-Proteobacteria, for example, in genomes of Enterobacteria, Vibrio, and Halomonas species, and in typical soil bacteria, such as Burkholderia, Herbaspirillum, and Rhizobium.

Sunlight-Triggered Nanoparticle Synergy: Teamwork of Reactive Oxygen Species and Nitric Oxide Released from Mesoporous Organosilica with Advanced Antibacterial Activity.

Colonization of surfaces by microorganisms is an urging problem. In combination with the increasing antibiotic resistance of pathogenic bacteria, severe infections are reported more frequently in medical settings. Therefore, there is a large demand to explore innovative surface coatings that provide intrinsic and highly effective antibacterial activity. Materials containing silver nanoparticles have been developed in the past for this purpose, but this solution has come into criticism due to various disadvantages like notable toxicity against higher organisms, the high price, and low abundance of silver. Here, we introduce a new, sunlight-mediated organosilica nanoparticle (NP) system based on silver-free antibacterial activity. The simultaneous release of nitric oxide (NO) in combination with singlet oxygen and superoxide radicals (O2(•-)) as reactive oxygen species (ROS) leads to the emergence of highly reactive peroxynitrite molecules with significantly enhanced biocidal activity. This special cooperative effect can only be realized, if the ROS-producing moieties and the functional entities releasing NO are spatially separated from each other. In one type of particle, Rose Bengal as an efficient singlet oxygen ((1)O2) producer was covalently bound to SH functionalities applying thiol-ene click chemistry. "Charging" the second type of particles with NO was realized by quantitatively transferring the thiol groups into S-nitrosothiol functionalities. We probed the oxidation power of ROS-NP alone and in combination with NO-NP using sunlight as a trigger. The high antibacterial efficiency of dual-action nanoparticles was demonstrated using disinfection assays with the pathogenic bacterium Pseudomonas aeruginosa.

Cloning, functional expression and characterization of a bifunctional 3-hydroxybutanal dehydrogenase /reductase involved in acetone metabolism by Desulfococcus biacutus.

BACKGROUND: The strictly anaerobic, sulfate-reducing bacterium Desulfococcus biacutus can utilize acetone as sole carbon and energy source for growth. Whereas in aerobic and nitrate-reducing bacteria acetone is activated by carboxylation with CO2 to acetoacetate, D. biacutus involves CO as a cosubstrate for acetone activation through a different, so far unknown pathway. Proteomic studies indicated that, among others, a predicted medium-chain dehydrogenase/reductase (MDR) superfamily, zinc-dependent alcohol dehydrogenase (locus tag DebiaDRAFT_04514) is specifically and highly produced during growth with acetone. RESULTS: The MDR gene DebiaDRAFT_04514 was cloned and overexpressed in E. coli. The purified recombinant protein required zinc as cofactor, and accepted NADH/NAD+ but not NADPH/NADP+ as electron donor/acceptor. The pH optimum was at pH 8, and the temperature optimum at 45 °C. Highest specific activities were observed for reduction of C3 - C5-aldehydes with NADH, such as propanal to propanol (380 ± 15 mU mg-1 protein), butanal to butanol (300 ± 24 mU mg-1), and 3-hydroxybutanal to 1,3-butanediol (248 ± 60 mU mg-1), however, the enzyme also oxidized 3-hydroxybutanal with NAD+ to acetoacetaldehyde (83 ± 18 mU mg-1). CONCLUSION: The enzyme might play a key role in acetone degradation by D. biacutus, for example as a bifunctional 3-hydroxybutanal dehydrogenase/reductase. Its recombinant production may represent an important step in the elucidation of the complete degradation pathway.

Bacteria may induce the secretion of mucin-like proteins by the diatom Phaeodactylum tricornutum.

Benthic diatoms live in photoautotrophic/heterotrophic biofilm communities embedded in a matrix of secreted extracellular polymeric substances. Closely associated bacteria influence their growth, aggregation, and secretion of exopolymers. We have studied a diatom/bacteria model community, in which a marine Roseobacter strain is able to grow with secreted diatom exopolymers as a sole source of carbon. The strain influences the aggregation of Phaeodactylum tricornutum by inducing a morphotypic transition from planktonic, fusiform cells to benthic, oval cells. Analysis of the extracellular soluble proteome of P. tricornutum in the presence and absence of bacteria revealed constitutively expressed newly identified proteins with mucin-like domains that appear to be typical for extracellular diatom proteins. In contrast to mucins, the proline-, serine-, threonine-rich (PST) domains in these proteins were also found in combination with protease-, glucosidase- and leucine-rich repeat-domains. Bioinformatic functional predictions indicate that several of these newly identified diatom-specific proteins may be involved in algal defense, intercellular signaling, and aggregation.

Acetone utilization by sulfate-reducing bacteria: draft genome sequence of Desulfococcus biacutus and a proteomic survey of acetone-inducible proteins.

BACKGROUND: The sulfate-reducing bacterium Desulfococcus biacutus is able to utilize acetone for growth by an inducible degradation pathway that involves a novel activation reaction for acetone with CO as a co-substrate. The mechanism, enzyme(s) and gene(s) involved in this acetone activation reaction are of great interest because they represent a novel and yet undefined type of activation reaction under strictly anoxic conditions. RESULTS: In this study, a draft genome sequence of D. biacutus was established. Sequencing, assembly and annotation resulted in 159 contigs with 5,242,029 base pairs and 4773 predicted genes; 4708 were predicted protein-encoding genes, and 3520 of these had a functional prediction. Proteins and genes were identified that are specifically induced during growth with acetone. A thiamine diphosphate-requiring enzyme appeared to be highly induced during growth with acetone and is probably involved in the activation reaction. Moreover, a coenzyme B12- dependent enzyme and proteins that are involved in redox reactions were also induced during growth with acetone. CONCLUSIONS: We present for the first time the genome of a sulfate reducer that is able to grow with acetone. The genome information of this organism represents an important tool for the elucidation of a novel reaction mechanism that is employed by a sulfate reducer in acetone activation.

Description of a 'plankton filtration bias' in sequencing-based bacterial community analysis and of an Arduino microcontroller-based flowmeter device that can help to resolve it.

Diversity studies of aquatic picoplankton (bacterioplankton) communities using size-class filtration, DNA extraction, PCR and sequencing of phylogenetic markers, require a robust methodological pipeline, since biases have been demonstrated essentially at all levels, including DNA extraction, primer choice and PCR. Even different filtration volumes of the same plankton sample and, thus, different biomass loading of the filters, can distort the sequencing results. In this study, we designed an Arduino microcontroller-based flowmeter that records the decrease of initial (maximal) flowrate as proxy for increasing biomass loading and clogging of filters during plankton filtration. The device was tested using freshwater plankton of Lake Constance, and total DNA was extracted and an 16S rDNA amplicon was sequenced. We confirmed that different filtration volumes used for the same water sample affect the sequencing results. Differences were visible in alpha and beta diversities and across all taxonomic ranks. Taxa most affected were typical freshwater Actinobacteria and Bacteroidetes, increasing up to 38% and decreasing up to 29% in relative abundance, respectively. In another experiment, a lake water sample was filtered undiluted and three-fold diluted, and each filtration was stopped once the flowrate had reduced to 50% of initial flowrate, hence, at the same degree of filter clogging. The three-fold diluted sample required three-fold filtration volumes, while equivalent amounts of total DNA were extracted and differences across all taxonomic ranks were not statistically significant compared to the undiluted controls. In conclusion, this work confirms a volume/biomass-dependent bacterioplankton filtration bias for sequencing-based community analyses and provides an improved procedure for controlling biomass loading during filtrations and recovery of equivalent amounts of DNA from samples independent of the plankton density. The application of the device can also avoid the distorting of sequencing results as caused by the plankton filtration bias.

Paracoccus denitrificans PD1222 utilizes hypotaurine via transamination followed by spontaneous desulfination to yield acetaldehyde and, finally, acetate for growth.

Hypotaurine (HT; 2-aminoethane-sulfinate) is known to be utilized by bacteria as a sole source of carbon, nitrogen, and energy for growth, as is taurine (2-aminoethane-sulfonate); however, the corresponding HT degradation pathway has remained undefined. Genome-sequenced Paracoccus denitrificans PD1222 utilized HT (and taurine) quantitatively for heterotrophic growth and released the HT sulfur as sulfite (and sulfate) and HT nitrogen as ammonium. Enzyme assays with cell extracts suggested that an HT-inducible HT:pyruvate aminotransferase (Hpa) catalyzes the deamination of HT in an initial reaction step. Partial purification of the Hpa activity and peptide fingerprinting-mass spectrometry (PF-MS) identified the Hpa candidate gene; it encoded an archetypal taurine:pyruvate aminotransferase (Tpa). The same gene product was identified via differential PAGE and PF-MS, as was the gene of a strongly HT-inducible aldehyde dehydrogenase (Adh). Both genes were overexpressed in Escherichia coli. The overexpressed, purified Hpa/Tpa showed HT:pyruvate-aminotransferase activity. Alanine, acetaldehyde, and sulfite were identified as the reaction products but not sulfinoacetaldehyde; the reaction of Hpa/Tpa with taurine yielded sulfoacetaldehyde, which is stable. The overexpressed, purified Adh oxidized the acetaldehyde generated during the Hpa reaction to acetate in an NAD(+)-dependent reaction. Based on these results, the following degradation pathway for HT in strain PD1222 can be depicted. The identified aminotransferase converts HT to sulfinoacetaldehyde, which desulfinates spontaneously to acetaldehyde and sulfite; the inducible aldehyde dehydrogenase oxidizes acetaldehyde to yield acetate, which is metabolized, and sulfite, which is excreted.

Biodegradable Mineral Plastics.

Mineral plastics are a promising class of bio-inspired materials that offer exceptional properties, like self-heal ability, stretchability in the hydrogel state, and high hardness, toughness, transparency, and non-flammability in the dry state along with reversible transformation into the hydrogel by addition of water. This enables easy reshape-ability and recycling like the solubility in mild acids to subsequently form mineral plastics again by base addition. However, current mineral plastics rely on petrochemistry, are hardly biodegradable, and thus persistent in nature. This work presents the next generation of mineral plastics, which are bio-based and biodegradable, making them a promising, new class of polymers for the development of environmentally friendly materials. Physically cross-linked (poly)glutamic-acid (PGlu)-based mineral plastics are synthesized using various alcohol-water mixtures, metal ion ratios and molecular weights. The rheological properties are easily adjusted using these parameters. The general procedure involves addition of equimolar solution of CaCl2 to PGlu in equal volumes followed by addition of iPrOH (iPrOH:H2 O = 1:1) under vigorous stirring conditions. The ready biodegradability of PGlu/CaFe mineral plastic is confirmed in this study where the elements N, Ca, and Fe present in it tend to act as additional nutrients, supporting the growth of microorganisms and consequently, promoting the biodegradation process.

Ecophysiology and interactions of a taurine-respiring bacterium in the mouse gut.

Taurine-respiring gut bacteria produce H2S with ambivalent impact on host health. We report the isolation and ecophysiological characterization of a taurine-respiring mouse gut bacterium. Taurinivorans muris strain LT0009 represents a new widespread species that differs from the human gut sulfidogen Bilophila wadsworthia in its sulfur metabolism pathways and host distribution. T. muris specializes in taurine respiration in vivo, seemingly unaffected by mouse diet and genotype, but is dependent on other bacteria for release of taurine from bile acids. Colonization of T. muris in gnotobiotic mice increased deconjugation of taurine-conjugated bile acids and transcriptional activity of a sulfur metabolism gene-encoding prophage in other commensals, and slightly decreased the abundance of Salmonella enterica, which showed reduced expression of galactonate catabolism genes. Re-analysis of metagenome data from a previous study further suggested that T. muris can contribute to protection against pathogens by the commensal mouse gut microbiota. Together, we show the realized physiological niche of a key murine gut sulfidogen and its interactions with selected gut microbiota members.

Two enzymes of a complete degradation pathway for linear alkylbenzenesulfonate (LAS) surfactants: 4-sulfoacetophenone Baeyer-Villiger monooxygenase and 4-sulfophenylacetate esterase in Comamonas testosteroni KF-1.

Complete biodegradation of the surfactant linear alkylbenzenesulfonate (LAS) is accomplished by complex bacterial communities in two steps. First, all LAS congeners are degraded into about 50 sulfophenylcarboxylates (SPC), one of which is 3-(4-sulfophenyl)butyrate (3-C(4)-SPC). Second, these SPCs are mineralized. 3-C(4)-SPC is mineralized by Comamonas testosteroni KF-1 in a process involving 4-sulfoacetophenone (SAP) as a metabolite and an unknown inducible Baeyer-Villiger monooxygenase (BVMO) to yield 4-sulfophenyl acetate (SPAc) from SAP (SAPMO enzyme); hydrolysis of SPAc to 4-sulfophenol and acetate is catalyzed by an unknown inducible esterase (SPAc esterase). Transcriptional analysis showed that one of four candidate genes for BVMOs in the genome of strain KF-1, as well as an SPAc esterase candidate gene directly upstream, was inducibly transcribed during growth with 3-C(4)-SPC. The same genes were identified by enzyme purification and peptide fingerprinting-mass spectrometry when SAPMO was enriched and SPAc esterase purified to homogeneity by protein chromatography. Heterologously overproduced pure SAPMO converted SAP to SPAc and was active with phenylacetone and 4-hydroxyacetophenone but not with cyclohexanone and progesterone. SAPMO showed the highest sequence homology to the archetypal phenylacetone BVMO (57%), followed by steroid BVMO (55%) and 4-hydroxyacetophenone BVMO (30%). Finally, the two pure enzymes added sequentially, SAPMO with NADPH and SAP, and then SPAc esterase, catalyzed the conversion of SAP via SPAc to 4-sulfophenol and acetate in a 1:1:1:1 molar ratio. Hence, the first two enzymes of a complete LAS degradation pathway were identified, giving evidence for the recruitment of members of the very versatile type I BVMO and carboxylester hydrolase enzyme families for the utilization of a xenobiotic compound by bacteria.