| biphenyl and benzoate metabolism in a genomic context: outlining genome-wide metabolic networks in burkholderia xenovorans lb400. | we designed and successfully implemented the use of in situ-synthesized 45-mer oligonucleotide dna microarrays (xeochips) for genome-wide expression profiling of burkholderia xenovorans lb400, which is among the best aerobic polychlorinated biphenyl degraders known so far. we conducted differential gene expression profiling during exponential growth on succinate, benzoate, and biphenyl as sole carbon sources and investigated the transcriptome of early-stationary-phase cells grown on biphenyl. ba ... | 2004 | 15294836 |
| evolution of the biphenyl dioxygenase bpha from burkholderia xenovorans lb400 by random mutagenesis of multiple sites in region iii. | it is now established that several amino acids of region iii of the biphenyl dioxygenase (bpdo) alpha subunit are involved in substrate recognition and regiospecificity toward chlorobiphenyls. however, the sequence pattern of the amino acids of that segment of seven amino acids located in the c-terminal portion of the alpha subunit is rather limited in bpdos of natural occurrence. in this work, we have randomly mutated simultaneously four residues (thr(335)-phe(336)-ile(338)-ile(341)) of region ... | 2004 | 15342624 |
| revisiting the regiospecificity of burkholderia xenovorans lb400 biphenyl dioxygenase toward 2,2'-dichlorobiphenyl and 2,3,2',3'-tetrachlorobiphenyl. | 2,2'-dichlorobiphenyl (cb) is transformed by the biphenyl dioxygenase of burkholderia xenovorans lb400 (lb400 bpdo) into two metabolites (1 and 2). the most abundant metabolite, 1, was previously identified as 2,3-dihydroxy-2'-chlorobiphenyl and was presumed to originate from the initial attack by the oxygenase on the chlorine-bearing ortho carbon and on its adjacent meta carbon of one phenyl ring. 2,3,2',3'-tetrachlorobiphenyl is transformed by lb400 bpdo into two metabolites that had never bee ... | 2004 | 15342625 |
| classification of the biphenyl- and polychlorinated biphenyl-degrading strain lb400t and relatives as burkholderia xenovorans sp. nov. | strain lb400t is the best-studied polychlorinated biphenyl (pcb) degrader. this organism has previously been allocated in the genus burkholderia, since its 16s rrna gene sequence shows 98.6 % sequence similarity to the type strains of burkholderia graminis and burkholderia terricola. a polyphasic study was undertaken to clarify the actual taxonomic position of this biotechnologically important organism and of two strains, one recovered from a blood culture vial and one from a coffee plant rhizos ... | 2004 | 15388727 |
| stereochemistry of the reaction catalysed by 2-hydroxy-6-keto-6-phenyl-hexa-2,4-dienoic acid 5,6-hydrolase (bphd). | the stereochemical course of the reaction catalysed by c-c hydrolase bphd from burkholderia xenovorans lb400 occurs with replacement of a benzoyl group by hydrogen with overall retention of stereochemistry. | 2004 | 15614396 |
| insight into pollutant bioavailability and toxicity using raman confocal microscopy. | raman confocal microscopy was used to discriminate between cultures of burkholderia xenovorans lb400 exposed to four different common environmental pollutants: phenanthrene, dodecane, 3-chlorobiphenyl and pentachlorophenol. evidence is presented for the application of raman spectroscopy as a bioassay for pollutant bioavailability and toxicity. | 2005 | 15649544 |
| metabolism of dibenzofuran and dibenzo-p-dioxin by the biphenyl dioxygenase of burkholderia xenovorans lb400 and comamonas testosteroni b-356. | we examined the metabolism of dibenzofuran (df) and dibenzo-p-dioxin (dd) by the biphenyl dioxygenase (bpdo) of comamonas testosteroni b-356 and compared it with that of burkholderia xenovorans lb400. data showed that both enzymes oxygenated df at a low rate, but escherichia coli cells expressing lb400 bpdo degraded df at higher rate (30 nmol in 18 h) compared with cells expressing b-356 bpdo (2 nmol in 18 h). furthermore, both bpdos produced dihydro-dihydroxy-dibenzofuran as a major metabolite, ... | 2005 | 15700128 |
| resolving the profile of metabolites generated during oxidation of dibenzofuran and chlorodibenzofurans by the biphenyl catabolic pathway enzymes. | although the metabolism of dibenzofuran by the biphenyl catabolic enzymes had been inferred in previous reports, the metabolic pattern has never been determined unambiguously. in this work, we describe the evolved biphenyl dioxygenase (bpdo) rr41 that exhibits a higher turnover rate of metabolism toward dibenzofuran and chlorodibenzofurans than the parental burkholderia xenovorans lb400 bpdo. we used rr41 bpdo to identify unambiguously the metabolites produced from the oxygenation of dibenzofura ... | 2005 | 16039530 |
| active-site engineering of biphenyl dioxygenase: effect of substituted amino acids on substrate specificity and regiospecificity. | biphenyl dioxygenase (bph dox) catalyzes the initial dioxygenation step in the metabolism of biphenyl. the large subunit (bpha1) of bph dox plays a crucial role in the determination of the substrate specificity of biphenyl-related compounds including polychlorinated biphenyls (pcbs). previously, the substitution of asn at thr-376 near the active-site iron in the bpha1 of pseudomonas pseudoalcaligenes kf707 expanded the oxidation range and altered the regiospecificity of bph dox for pcbs. in this ... | 2006 | 16217654 |
| growth substrate- and phase-specific expression of biphenyl, benzoate, and c1 metabolic pathways in burkholderia xenovorans lb400. | recent microarray experiments suggested that burkholderia xenovorans lb400, a potent polychlorinated biphenyl (pcb)-degrading bacterium, utilizes up to three apparently redundant benzoate pathways and a c(1) metabolic pathway during biphenyl and benzoate metabolism. to better characterize the roles of these pathways, we performed quantitative proteome profiling of cells grown on succinate, benzoate, or biphenyl and harvested during either mid-logarithmic growth or the transition between the loga ... | 2005 | 16291673 |
| biphenyl degradation kinetics by burkholderia xenovorans lb400 in two-phase partitioning bioreactors. | biphenyl could be successfully degraded by burkholderia xenovorans lb400, initially described as pseudomonas sp. lb400, in two-phase partitioning bioreactors (tppbs). tppbs are comprised of an aqueous, cell containing phase, and an immiscible, biocompatible organic phase that partitions toxic and/or poorly soluble substrates (in this case biphenyl) based on maintaining a thermodynamic equilibrium. the critical logko/w of the organism was found to be approximately 5.5, indicating that solvents wi ... | 2006 | 16310831 |
| genetic and genomic insights into the role of benzoate-catabolic pathway redundancy in burkholderia xenovorans lb400. | transcriptomic and proteomic analyses of burkholderia xenovorans lb400, a potent polychlorinated biphenyl (pcb) degrader, have implicated growth substrate- and phase-dependent expression of three benzoate-catabolizing pathways: a catechol ortho cleavage (ben-cat) pathway and two benzoyl-coenzyme a pathways, encoded by gene clusters on the large chromosome (boxc) and the megaplasmid (boxm). to elucidate the significance of this apparent redundancy, we constructed mutants with deletions of the ben ... | 2006 | 16391095 |
| the clc element of pseudomonas sp. strain b13, a genomic island with various catabolic properties. | pseudomonas sp. strain b13 is a bacterium known to degrade chloroaromatic compounds. the properties to use 3- and 4-chlorocatechol are determined by a self-transferable dna element, the clc element, which normally resides at two locations in the cell's chromosome. here we report the complete nucleotide sequence of the clc element, demonstrating the unique catabolic properties while showing its relatedness to genomic islands and integrative and conjugative elements rather than to other known cata ... | 2006 | 16484212 |
| generation of novel-substrate-accepting biphenyl dioxygenases through segmental random mutagenesis and identification of residues involved in enzyme specificity. | aryl-hydroxylating dioxygenases are of interest for the degradation of persistant aromatic pollutants, such as polychlorobiphenyls (pcbs), or as catalysts for the functionalization of aromatic scaffolds. in order to achieve dioxygenation of technical mixtures of pcbs, enzymes with broadened or altered substrate ranges are essential. to alter the substrate specificity of the biphenyl dioxygenase (bpha) of burkholderia xenovorans lb400, we applied a directed evolution approach that used structure- ... | 2006 | 16517671 |
| nitrate-dependent anaerobic carbon monoxide oxidation by aerobic co-oxidizing bacteria. | two dissimilatory nitrate-reducing (burkholderia xenovorans lb400 and xanthobacter sp. str. cox) and two denitrifying isolates (stappia aggregata iam 12614 and bradyrhizobium sp. str. cpp), previously characterized as aerobic co oxidizers, consumed co at ecologically relevant levels (<100 ppm) under anaerobic conditions in the presence, but not absence, of nitrate. none of the isolates were able to grow anaerobically with co as a carbon or energy source, however, and nitrate-dependent anaerobic ... | 2006 | 16542399 |
| polychlorinated biphenyl (pcb)-degrading bacteria associated with trees in a pcb-contaminated site. | the abundance, identities, and degradation abilities of indigenous polychlorinated biphenyl (pcb)-degrading bacteria associated with five species of mature trees growing naturally in a contaminated site were investigated to identify plants that enhance the microbial pcb degradation potential in soil. culturable pcb degraders were associated with every plant species examined in both the rhizosphere and root zone, which was defined as the bulk soil in which the plant was rooted. significantly high ... | 2006 | 16597927 |
| degradation of aroclor 1242 dechlorination products in sediments by burkholderia xenovorans lb400(ohb) and rhodococcus sp. strain rha1(fcb). | burkholderia xenovorans strain lb400, which possesses the biphenyl pathway, was engineered to contain the oxygenolytic ortho dehalogenation (ohb) operon, allowing it to grow on 2-chlorobenzoate and to completely mineralize 2-chlorobiphenyl. a two-stage anaerobic/aerobic biotreatment process for aroclor 1242-contaminated sediment was simulated, and the degradation activities and genetic stabilities of lb400(ohb) and the previously constructed strain rha1(fcb), capable of growth on 4-chlorobenzoat ... | 2006 | 16597946 |
| chlorophenol hydroxylases encoded by plasmid pjp4 differentially contribute to chlorophenoxyacetic acid degradation. | phenoxyalkanoic compounds are used worldwide as herbicides. cupriavidus necator jmp134(pjp4) catabolizes 2,4-dichlorophenoxyacetate (2,4-d) and 4-chloro-2-methylphenoxyacetate (mcpa), using tfd functions carried on plasmid pjp4. tfda cleaves the ether bonds of these herbicides to produce 2,4-dichlorophenol (2,4-dcp) and 4-chloro-2-methylphenol (mcp), respectively. these intermediates can be degraded by two chlorophenol hydroxylases encoded by the tfdb(i) and tfdb(ii) genes to produce the respect ... | 2006 | 16597983 |
| a glutathione s-transferase catalyzes the dehalogenation of inhibitory metabolites of polychlorinated biphenyls. | bphk is a glutathione s-transferase of unclear physiological function that occurs in some bacterial biphenyl catabolic (bph) pathways. we demonstrated that bphk of burkholderia xenovorans strain lb400 catalyzes the dehalogenation of 3-chloro 2-hydroxy-6-oxo-6-phenyl-2,4-dienoates (hopdas), compounds that are produced by the cometabolism of polychlorinated biphenyls (pcbs) by the bph pathway and that inhibit the pathway's hydrolase. a one-column protocol was developed to purify heterologously pro ... | 2006 | 16740949 |
| structures of ternary complexes of bphk, a bacterial glutathione s-transferase that reductively dechlorinates polychlorinated biphenyl metabolites. | prokaryotic glutathione s-transferases are as diverse as their eukaryotic counterparts but are much less well characterized. bphk from burkholderia xenovorans lb400 consumes two gsh molecules to reductively dehalogenate chlorinated 2-hydroxy-6-oxo-6-phenyl-2,4-dienoates (hopdas), inhibitory polychlorinated biphenyl metabolites. crystallographic structures of two ternary complexes of bphk were solved to a resolution of 2.1a. in the bphk-gsh-hopda complex, gsh and hopda molecules occupy the g- and ... | 2006 | 16920719 |
| kinetic and structural insight into the mechanism of bphd, a c-c bond hydrolase from the biphenyl degradation pathway. | kinetic and structural analyses of 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid (hopda) hydrolase from burkholderia xenovorans lb400 (bphd(lb400)) provide insight into the catalytic mechanism of this unusual serine hydrolase. single turnover stopped-flow analysis at 25 degrees c showed that the enzyme rapidly (1/tau(1) approximately 500 s(-1)) transforms hopda (lambda(max) = 434 nm) into a species with electronic absorption maxima at 473 and 492 nm. the absorbance of this enzyme-bound species ( ... | 2006 | 16964968 |
| expression of bacterial biphenyl-chlorobiphenyl dioxygenase genes in tobacco plants. | optimized plant-microbe bioremediation processes in which the plant initiates the metabolism of xenobiotics and releases the metabolites in the rhizosphere to be further degraded by the rhizobacteria is a promising alternative to restore contaminated sites in situ. however, such processes require that plants produce the metabolites that bacteria can readily oxidize. the biphenyl dioxygenase is the first enzyme of the bacterial catabolic pathway involved in the degradation of polychlorinated biph ... | 2007 | 17006888 |
| coping with polychlorinated biphenyl (pcb) toxicity: physiological and genome-wide responses of burkholderia xenovorans lb400 to pcb-mediated stress. | the biodegradation of polychlorinated biphenyls (pcbs) relies on the ability of aerobic microorganisms such as burkholderia xenovorans sp. lb400 to tolerate two potential modes of toxicity presented by pcb degradation: passive toxicity, as hydrophobic pcbs potentially disrupt membrane and protein function, and degradation-dependent toxicity from intermediates of incomplete degradation. we monitored the physiological characteristics and genome-wide expression patterns of lb400 in response to the ... | 2006 | 17021212 |
| evidence for a gem-diol reaction intermediate in bacterial c-c hydrolase enzymes bphd and mhpc from 13c nmr spectroscopy. | c-c hydrolase enzymes mhpc and bphd catalyze the hydrolytic c-c cleavage of meta-ring fission intermediates on the escherichia coli phenylpropionic acid and burkholderia xenovorans lb400 biphenyl degradation pathways and are both members of the alpha/beta-hydrolase family containing a ser-his-asp catalytic triad. the catalytic mechanism of this family of enzymes is thought to proceed via a gem-diol reaction intermediate, which has not been observed directly. site-directed single mutants of bphd ... | 2006 | 17029401 |
| catalytic role for arginine 188 in the c-c hydrolase catalytic mechanism for escherichia coli mhpc and burkholderia xenovorans lb400 bphd. | the alpha/beta-hydrolase superfamily, comprised mainly of esterase and lipase enzymes, contains a family of bacterial c-c hydrolases, including mhpc and bphd which catalyze the hydrolytic c-c cleavage of meta-ring fission intermediates on the escherichia coli phenylpropionic acid pathway and burkholderia xenovorans lb400 biphenyl degradation pathway, respectively. five active site amino acid residues (arg-188, asn-109, phe-173, cys-261, and trp-264) were identified from sequence alignments that ... | 2006 | 17029402 |
| the complete genome of rhodococcus sp. rha1 provides insights into a catabolic powerhouse. | rhodococcus sp. rha1 (rha1) is a potent polychlorinated biphenyl-degrading soil actinomycete that catabolizes a wide range of compounds and represents a genus of considerable industrial interest. rha1 has one of the largest bacterial genomes sequenced to date, comprising 9,702,737 bp (67% g+c) arranged in a linear chromosome and three linear plasmids. a targeted insertion methodology was developed to determine the telomeric sequences. rha1's 9,145 predicted protein-encoding genes are exceptional ... | 2006 | 17030794 |
| burkholderia xenovorans lb400 harbors a multi-replicon, 9.73-mbp genome shaped for versatility. | burkholderia xenovorans lb400 (lb400), a well studied, effective polychlorinated biphenyl-degrader, has one of the two largest known bacterial genomes and is the first nonpathogenic burkholderia isolate sequenced. from an evolutionary perspective, we find significant differences in functional specialization between the three replicons of lb400, as well as a more relaxed selective pressure for genes located on the two smaller vs. the largest replicon. high genomic plasticity, diversity, and speci ... | 2006 | 17030797 |
| burkholderia ginsengisoli sp. nov., a beta-glucosidase-producing bacterium isolated from soil of a ginseng field. | a bacterial strain (designated kmy03t) that possesses beta-glucosidase activity was isolated from soil from a ginseng field in south korea and was characterized in order to determine its taxonomic position. the bacterium was found to comprise gram-negative, rod-shaped, motile cells with unipolar polytrichous flagella. on the basis of 16s rrna gene sequence similarity, strain kmy03t was shown to belong to the family burkholderiaceae of the betaproteobacteria, being most closely related to burkhol ... | 2006 | 17082385 |
| family shuffling of soil dna to change the regiospecificity of burkholderia xenovorans lb400 biphenyl dioxygenase. | previous work has shown that the c-terminal portion of bpha, especially two amino acid segments designated region iii and region iv, influence the regiospecificity of the biphenyl dioxygenase (bpdo) toward 2,2'-dichlorobiphenyl (2,2'-cb). in this work, we evolved bpdo by shuffling bpha genes amplified from polychlorinated biphenyl-contaminated soil dna. sets of approximately 1-kb dna fragments were amplified with degenerate primers designed to amplify the c-terminal portion of bpha. these fragme ... | 2007 | 17142386 |
| response to (chloro)biphenyls of the polychlorobiphenyl-degrader burkholderia xenovorans lb400 involves stress proteins also induced by heat shock and oxidative stress. | we report the effects of 4-chlorobiphenyl and biphenyl on the physiology, morphology and proteome of the polychlorobiphenyl-degrader burkholderia xenovorans lb400. the exposure to 4-chlorobiphenyl decreases the growth of lb400 on glucose, and cells exhibit irregular outer membranes, a larger periplasmic space and electron-dense granules in the cytoplasm. additionally, lysis of cells was observed during incubation with 4-chlorobiphenyl or biphenyl. proteome of b. xenovorans lb400 exposed to biphe ... | 2007 | 17166226 |
| investigation of a general base mechanism for ester hydrolysis in c-c hydrolase enzymes of the alpha/beta-hydrolase superfamily: a novel mechanism for the serine catalytic triad. | previous mechanistic and crystallographic studies on two c-c hydrolase enzymes, escherichia coli mhpc and burkholderia xenovorans bphd, support a general base mechanism for c-c hydrolytic cleavage, rather than the nucleophilic mechanism expected for a serine hydrolase. the role of the active site serine residue could be to form a hydrogen bond with a gem-diolate intermediate, or to protonate such an intermediate. hydrolase bphd is able to catalyse the hydrolysis of p-nitrophenyl benzoate ester s ... | 2007 | 17252134 |
| generation by a widely applicable approach of a hybrid dioxygenase showing improved oxidation of polychlorobiphenyls. | recently, a sequence-based approach has been developed for the fast isolation and characterization of class ii aryl-hydroxylating dioxygenase activities (s. kahl and b. hofer, microbiology 149:1475-1481, 2003). it comprises the pcr amplification of segments of alpha subunit genes of unknown sequence that encode the catalytic center and their fusion with sequences of the bpha gene cluster of burkholderia xenovorans lb400. one of the resulting chimeric enzymes, harboring the core segment of a diox ... | 2007 | 17322323 |
| motility and chemotaxis of pseudomonas sp. b4 towards polychlorobiphenyls and chlorobenzoates. | the polychlorinated biphenyl (pcb)-degrading pseudomonas sp. b4 was tested for its motility and ability to sense and respond to biphenyl, its chloroderivatives and chlorobenzoates in chemotaxis assays. pseudomonas sp. b4 was attracted to biphenyl, pcbs and benzoate in swarm plate and capillary assays. chemotaxis towards these compounds correlated with their use as carbon and energy sources. no chemotactic effect was observed in the presence of 2- and 3-chlorobenzoates. furthermore, a toxic effec ... | 2007 | 17374130 |
| synthesis of highly hydroxylated aromatics by evolved biphenyl dioxygenase and subsequent dihydrodiol dehydrogenase. | the evolved bpha1 (2049) gene, in which nine amino acids from the pseudomonas pseudoalcaligenes kf707 bpha1 were changed to those from the burkholderia xenovorans lb400 bpha1 (m247i, h255q, v258i, g268a, d303e, -313g, s324t, v325i, and t376n), was expressed in escherichia coli along with the bpha2a3a4 and bphb genes derived from strain kf707. this recombinant e. coli cells converted biphenyl and several heterocyclic aromatic compounds into the highly hydroxylated products such as biphenyl-2,3,2' ... | 2007 | 17401562 |
| characterization of a c-c bond hydrolase from sphingomonas wittichii rw1 with novel specificities towards polychlorinated biphenyl metabolites. | sphingomonas wittichii rw1 degrades chlorinated dibenzofurans and dibenzo-p-dioxins via meta cleavage. we used inverse pcr to amplify dxnb2, a gene encoding one of three meta-cleavage product (mcp) hydrolases identified in the organism that are homologues of bphd involved in biphenyl catabolism. purified dxnb2 catalyzed the hydrolysis of 8-oh 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoate (hopda) approximately six times faster than for hopda at saturating substrate concentrations. moreover, the speci ... | 2007 | 17416660 |
| the tautomeric half-reaction of bphd, a c-c bond hydrolase. kinetic and structural evidence supporting a key role for histidine 265 of the catalytic triad. | bphd of burkholderia xenovorans lb400 catalyzes an unusual c-c bond hydrolysis of 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid (hopda) to afford benzoic acid and 2-hydroxy-2,4-pentadienoic acid (hpd). an enol-keto tautomerization has been proposed to precede hydrolysis via a gem-diol intermediate. the role of the canonical catalytic triad (ser-112, his-265, asp-237) in mediating these two half-reactions remains unclear. we previously reported that the bphd-catalyzed hydrolysis of hopda (lambda( ... | 2007 | 17442675 |
| chlorobenzoate inhibits growth and induces stress proteins in the pcb-degrading bacterium burkholderia xenovorans lb400. | aerobic bacteria, such as burkholderia xenovorans lb400, are able to degrade a wide range of polychlorobiphenyls (pcbs). generally, these bacteria are not able to transform chlorobenzoates (cbas), which accumulate during pcb degradation. in this study, the effects of cbas on the growth, the morphology and the proteome of burkholderia xenovorans lb400 were analysed. 4-cba and 2-cba were observed to inhibit the growth of strain lb400 on glucose. strain lb400 exposed to 4-cba exhibited increased nu ... | 2007 | 17522847 |
| characterization of biphenyl dioxygenase of pandoraea pnomenusa b-356 as a potent polychlorinated biphenyl-degrading enzyme. | biphenyl dioxygenase (bpdo) catalyzes the aerobic transformation of biphenyl and various polychlorinated biphenyls (pcbs). in three different assays, bpdo(b356) from pandoraea pnomenusa b-356 was a more potent pcb-degrading enzyme than bpdo(lb400) from burkholderia xenovorans lb400 (75% amino acid sequence identity), transforming nine congeners in the following order of preference: 2,3',4-trichloro approximately 2,3,4'-trichloro > 3,3'-dichloro > 2,4,4'-trichloro > 4,4'-dichloro approximately 2, ... | 2007 | 17526697 |
| polymer selection for biphenyl degradation in a solid-liquid two-phase partitioning bioreactor. | the commercially available thermoplastic polymer hytrel was selected as the delivery phase for the hydrophobic model compound biphenyl in a solid-liquid two-phase partitioning bioreactor (tppb), and 2.9 g biphenyl could successfully be degraded in 1-l tppbs by a pure culture of the biphenyl-degrading bacterium burkholderia xenovorans lb400 in 50 h and by a mixed microbial consortium isolated from contaminated soil in 45 h. tppbs consist of an aqueous cell-containing phase and an immiscible secon ... | 2007 | 17559230 |
| a large gene cluster in burkholderia xenovorans encoding abietane diterpenoid catabolism. | abietane diterpenoids are defense compounds synthesized by trees that are abundant in natural environments and occur as significant pollutants from pulp and paper production. burkholderia xenovorans lb400 has diverse catabolic capabilities and represents an important group of heterotrophic bacteria in soil environments. the genome sequence of lb400 revealed homologs of the dit genes of pseudomonas abietaniphila bkme-9, which encode abietane diterpenoid degradation. lb400 grew on abietic acid (ab ... | 2007 | 17586638 |
| enhancement of pcb degradation by burkholderia xenovorans lb400 in biphasic systems by manipulating culture conditions. | two-phase partitioning bioreactors (tppbs) can be used to biodegrade environmental contaminants after their extraction from soil. tppbs are typically stirred tank bioreactors containing an aqueous phase hosting the degrading microorganism and an immiscible, non-toxic and non-bioavailable organic phase functioning as a reservoir for hydrophobic compounds. biodegradation of these compounds in the aqueous phase results in thermodynamic disequilibrium and partitioning of additional compounds from th ... | 2008 | 17705226 |
| biochemical and structural characterization of the paralogous benzoate coa ligases from burkholderia xenovorans lb400: defining the entry point into the novel benzoate oxidation (box) pathway. | xenobiotic aromatic compounds represent one of the most significant classes of environmental pollutants. a novel benzoate oxidation (box) pathway has been identified recently in burkholderia xenovorans lb400 (referred to simply as lb400) that is capable of assimilating benzoate and intimately tied to the degradation of polychlorinated biphenyls (pcbs). the box pathway in lb400 is present in two paralogous copies (boxm and boxc) and encodes eight enzymes with the first committed step catalyzed by ... | 2007 | 17884091 |
| biodegradation of pcbs in two-phase partitioning bioreactors following solid extraction from soil. | this article demonstrates the feasibility of a novel process concept for the remediation of pcb contaminated soil. the proposed process consists of pcb extraction from soil using solid polymer beads, followed by biodegradation of the extracted pcbs in a solid-liquid two-phase partitioning bioreactor (tppb), where pcbs are delivered from the polymer beads to the degrading organisms. the commercially available thermoplastic polymer hytrel was used to extract aroclor 1242 from contaminated artifici ... | 2008 | 17929320 |
| distinct roles for two cyp226 family cytochromes p450 in abietane diterpenoid catabolism by burkholderia xenovorans lb400. | the 80-kb dit cluster of burkholderia xenovorans lb400 encodes the catabolism of abietane diterpenoids. this cluster includes ditq and ditu, predicted to encode cytochromes p450 (p450s) belonging to the poorly characterized cyp226a subfamily. using proteomics, we identified 16 dit-encoded proteins that were significantly more abundant in lb400 cells grown on dehydroabietic acid (dha) or abietic acid (aba) than in succinate-grown cells. a key difference in the catabolism of dha and aba lies in th ... | 2008 | 18156276 |
| burkholderia sediminicola sp. nov., isolated from freshwater sediment. | a gram-negative, motile and rod-shaped bacterium, designated hu2-65w(t), was isolated from freshwater sediment. the strain possessed ubiquinone 8 as the predominant isoprenoid quinone and contained major amounts of omega7-cis-octadecenoic acid and hexadecanoic acid in its cell envelope, which are properties shared by members of the genus burkholderia. on the basis of 16s rrna gene sequence similarity, strain hu2-65w(t) was most closely related to the type strain of burkholderia xenovorans (98.4 ... | 2008 | 18319455 |
| biphenyl dioxygenase from an arctic isolate is not cold adapted. | biphenyl dioxygenase from the psychrotolerant bacterium pseudomonas sp. strain cam-1 (bpdo(cam-1)) was purified and found to have an apparent k(cat) for biphenyl of 1.1 +/- 0.1 s(-1) (mean +/- standard deviation) at 4 degrees c. in contrast, bpdo(lb400) from the mesophile burkholderia xenovorans lb400 had no detectable activity at this temperature. at 57 degrees c, the half-life of the bpdo(cam-1) oxygenase was less than half that of the bpdo(lb400) oxygenase. nevertheless, bpdo(cam-1) appears t ... | 2008 | 18424535 |
| purification, crystallization and x-ray diffraction analysis of a novel ring-cleaving enzyme (boxc(c)) from burkholderia xenovorans lb400. | the assimilation of aromatic compounds by microbial species requires specialized enzymes to cleave the thermodynamically stable ring. in the recently discovered benzoate-oxidation (box) pathway in burkholderia xenovorans lb400, this is accomplished by a novel dihydrodiol lyase (boxc(c)). sequence analysis suggests that boxc(c) is part of the crotonase superfamily but includes an additional uncharacterized region of approximately 115 residues that is predicted to mediate ring cleavage. processing ... | 2008 | 18453716 |
| structural and biochemical characterization of a novel aldehyde dehydrogenase encoded by the benzoate oxidation pathway in burkholderia xenovorans lb400. | the recently identified benzoate oxidation (box) pathway in burkholderia xenovorans lb400 (lb400 hereinafter) assimilates benzoate through a unique mechanism where each intermediate is processed as a coenzyme a (coa) thioester. a key step in this process is the conversion of 3,4-dehydroadipyl-coa semialdehyde into its corresponding coa acid by a novel aldehyde dehydrogenase (aldh) (ec 1.2.1.x). the goal of this study is to characterize the biochemical and structural properties of the chromosomal ... | 2008 | 18462753 |
| the transcription regulator rcom-2 from burkholderia xenovorans is a cysteine-ligated hemoprotein that undergoes a redox-mediated ligand switch. | spectroscopic characterization of the newly discovered heme-pas domain sensor protein bxrcom-2 reveals that this protein undergoes redox-dependent ligand switching and co- and no-induced ligand displacement. the aerobic bacterium burkholderia xenovorans expresses two homologous heme-containing proteins that promote co-dependent transcription in vivo. these regulators of co metabolism, bxrcom-1 and bxrcom-2, are gas-responsive heme-pas domain proteins like mammalian neuronal pas domain protein 2 ... | 2008 | 18672900 |
| bacterial metabolism of polychlorinated biphenyls. | microbial metabolism is responsible for the removal of persistent organic pollutants including pcbs from the environment. anaerobic dehalogenation of highly chlorinated congeners in aquatic sediments is an important process, and recent evidence has indicated that dehalococcoides and related organisms are predominantly responsible for this process. such anaerobic dehalogenation generates lower chlorinated congeners which are easily degraded aerobically by enzymes of the biphenyl upper pathway (bp ... | 2008 | 18685266 |
| diversity of the c-terminal portion of the biphenyl dioxygenase large subunit. | the biphenyl dioxygenase (bpdo) catalyses a stereospecific dioxygenation of biphenyl and analogs of it. aside from being involved in the destruction and detoxification of toxic pollutants in soil, in the context of the green chemistry concept, this enzyme is a promising biocatalyst to design new more selective and more environmentally friendly approaches to manufacture fine chemicals. at this time, most of our knowledge about the variability of key residues determining the substrate specificity ... | 2008 | 18685267 |
| high rate of n2 fixation by east siberian cryophilic soil bacteria as determined by measuring acetylene reduction in nitrogen-poor medium solidified with gellan gum. | for evaluating n(2) fixation of diazotrophic bacteria, nitrogen-poor liquid media supplemented with at least 0.5% sugar and 0.2% agar are widely used for acetylene reduction assays. in such a soft gel medium, however, many n(2)-fixing soil bacteria generally show only trace acetylene reduction activity. here, we report that use of a n(2) fixation medium solidified with gellan gum instead of agar promoted growth of some gellan-preferring soil bacteria. in a soft gel medium solidified with 0.3% ge ... | 2009 | 19286791 |
| structural and biophysical characterization of boxc from burkholderia xenovorans lb400: a novel ring-cleaving enzyme in the crotonase superfamily. | the mineralization of aromatic compounds by microorganisms relies on a structurally and functionally diverse group of ring-cleaving enzymes. the recently discovered benzoate oxidation pathway in burkholderia xenovorans lb400 encodes a novel such ring-cleaving enzyme, termed boxc, that catalyzes the conversion of 2,3-dihydro-2,3-dihydroxybenzoyl-coa to 3,4-dehydroadipyl-coa without the requirement for molecular oxygen. sequence analysis indicates that boxc is a highly divergent member of the crot ... | 2009 | 19369256 |
| new mobile genetic elements in cupriavidus metallidurans ch34, their possible roles and occurrence in other bacteria. | cupriavidus metallidurans strain ch34 is a beta-proteobacterium that thrives in low concentrations of heavy metals. the genetic determinants of resistance to heavy metals are located on its two chromosomes, and are particularly abundant in the two megaplasmids, pmol28 and pmol30. we explored the involvement of mobile genetic elements in acquiring these and others traits that might be advantageous in this strain using genome comparison of cupriavidus/ralstonia strains and related beta-proteobacte ... | 2009 | 19390985 |
| identification of a novel alkaliphilic esterase active at low temperatures by screening a metagenomic library from antarctic desert soil. | a novel esterase was identified through functional screening of a metagenomic library in escherichia coli obtained from antarctic desert soil. the 297-amino-acid sequence had only low (<29%) similarity to a putative esterase from burkholderia xenovorans. the enzyme was active over a temperature range of 7 to 54 degrees c and at alkaline ph levels and is a potential candidate for industrial application. | 2009 | 19411411 |
| characterization of an aldolase-dehydrogenase complex that exhibits substrate channeling in the polychlorinated biphenyls degradation pathway. | an aldolase and dehydrogenase complex from the polychlorinated biphenyl degradation pathway of the bacterium burkholderia xenovorans lb400 was purified. the aldolase, bphi, had the highest activity with mn(2+) as the cofactor and was able to transform 4-hydroxy-2-oxopentanoate and 4-hydroxy-2-oxohexanoate to pyruvate and acetaldehyde or propionaldehyde with similar specificity constants. aldolase activity was competitively inhibited by the pyruvate enolate analogue, oxalate, with a k(ic) of 0.93 ... | 2009 | 19476337 |
| investigative mining of sequence data for novel enzymes: a case study with nitrilases. | mining sequence data is increasingly important for biocatalysis research. however, when relying on sequence data alone, prediction of the reaction catalyzed by a specific protein sequence is often elusive, and substrate specificity is far from trivial. the present study demonstrated an approach of combining sequence data and structures from distant homologs to target identification of new nitrilases that specifically utilize hindered nitrile substrates like mandelonitrile. a total of 212 non-ide ... | 2009 | 19539670 |
| environmentally relevant parameters affecting pcb degradation: carbon source- and growth phase-mitigated effects of the expression of the biphenyl pathway and associated genes in burkholderia xenovorans lb400. | the principal means for microbial degradation of polychlorinated biphenyls (pcbs) is through the biphenyl pathway. although molecular aspects of the regulation of the biphenyl pathway have been studied, information on environmental facets such as the effect of alternative carbon sources on (polychlorinated) biphenyl degradation is limited. here we explore the effect of environmental conditions (e.g., carbon source and growth phase) on the variation in pcb degradation profiles of burkholderia xen ... | 2010 | 19672561 |
| aerobic biotransformation of polybrominated diphenyl ethers (pbdes) by bacterial isolates. | polybrominated diphenyl ethers (pbdes) are flame retardants that have been used in consumer products and furniture for three decades. currently, very little is known about their fate in the environment and specifically about their susceptibility to aerobic biotransformation. here, we investigated the ability of the polychlorinated biphenyl (pcb) degrading bacteria rhodococcus jostii rha1 and burkholderia xenovorans lb400 to transform mono- through hexa-bdes at ppb levels. we also tested the pbde ... | 2009 | 19731666 |
| functional characterization of an orphan cupin protein from burkholderia xenovorans reveals a mononuclear nonheme fe2+-dependent oxygenase that cleaves beta-diketones. | cupins constitute a large and widespread superfamily of beta-barrel proteins in which a mononuclear metal site is both a conserved feature of the structure and a source of functional diversity. metal-binding residues are contributed from two core motifs that provide the signature for the superfamily. on the basis of conservation of this two-motif structure, we have identified an orf in the genome of burkholderia xenovorans that encodes a novel cupin protein (bxe_a2876) of unknown function. recom ... | 2009 | 19754880 |
| terminal-restriction fragment length polymorphism analysis of biphenyl dioxygenase genes from a polychlorinated biphenyl-polluted soil. | polychlorinated biphenyls (pcbs) are ubiquitous persistent organic pollutants that can be co-metabolically biotransformed by biphenyl-utilizing bacteria. in this study, terminal-restriction fragment length polymorphism (t-rflp) was applied to the substrate specificity-determining region of the 2,3-biphenyl dioxygenase encoding genes of a microbial community found in a pcb-polluted soil. notably, both the total biphenyl/pcb-utilizing community and its members actively expressing the 2,3-biphenyl ... | 2009 | 19835952 |
| purification and crystallization of a putative transcriptional regulator of the benzoate oxidation pathway in burkholderia xenovorans lb400. | burkholderia xenovorans lb400 harbours two paralogous copies of the recently discovered benzoate oxidation (box) pathway. while both copies are functional, the paralogues are differentially regulated and flanked by putative transcriptional regulators from distinct families. the putative lysr-type transcriptional regulator (lttr) adjacent to the megaplasmid-encoded box enzymes, bxe_c0898, has been produced recombinantly in escherichia coli and purified to homogeneity. gel-filtration studies show ... | 2009 | 19851006 |
| icmf is a fusion between the radical b12 enzyme isobutyryl-coa mutase and its g-protein chaperone. | coenzyme b(12) is used by two highly similar radical enzymes, which catalyze carbon skeleton rearrangements, methylmalonyl-coa mutase and isobutyryl-coa mutase (icm). icm catalyzes the reversible interconversion of isobutyryl-coa and n-butyryl-coa and exists as a heterotetramer. in this study, we have identified >70 bacterial proteins, which represent fusions between the subunits of icm and a p-loop gtpase and are currently misannotated as methylmalonyl-coa mutases. we designate this fusion prot ... | 2010 | 19864421 |
| mineralization of pcbs by the genetically modified strain cupriavidus necator jms34 and its application for bioremediation of pcbs in soil. | polychlorobiphenyls (pcbs) are classified as "high-priority pollutants." diverse microorganisms are able to degrade pcbs. however, bacterial degradation of pcbs is generally incomplete, leading to the accumulation of chlorobenzoates (cbas) as dead-end metabolites. to obtain a microorganism able to mineralize pcb congeners, the bph locus of burkholderia xenovorans lb400, which encodes one of the most effective pcb degradation pathways, was incorporated into the genome of the cba-degrading bacteri ... | 2010 | 20414654 |
| hybrid pseudomonads engineered by two-step homologous recombination acquire novel degradation abilities toward aromatics and polychlorinated biphenyls. | pseudomonas pseudoalcaligenes kf707 possesses a chromosomally encoded bph gene cluster responsible for the catabolism of biphenyl and polychlorinated biphenyls. previously, we constructed chimeric versions of the bpha1 gene, which encodes a large subunit of biphenyl dioxygenase, by using dna shuffling between bpha1 genes from p. pseudoalcaligenes kf707 and burkholderia xenovorans lb400. in this study, we demonstrate replacement of the bpha1 gene with chimeric bpha1 sequence within the chromosoma ... | 2010 | 20809076 |
| biodegradation of polyfluorinated biphenyl in bacteria. | fluorinated aromatic compounds are significant environmental pollutants, and microorganisms play important roles in their biodegradation. the effect of fluorine substitution on the transformation of fluorobiphenyl in two bacteria was investigated. pseudomonas pseudoalcaligenes kf707 and burkholderia xenovorans lb400 used 2,3,4,5,6-pentafluorobiphenyl and 4,4'-difluorobiphenyl as sole sources of carbon and energy. the catabolism of the fluorinated compounds was examined by gas chromatography-mass ... | 2010 | 20830605 |
| triclosan susceptibility and co-metabolism--a comparison for three aerobic pollutant-degrading bacteria. | the antimicrobial agent triclosan is an emerging and persistent environmental pollutant. this study evaluated the susceptibility and biodegradation potential of triclosan by three bacterial strains (sphingomonas wittichii rw1, burkholderia xenovorans lb400 and sphingomonas sp. ph-07) that are able to degrade aromatic pollutants (dibenzofuran, biphenyl and diphenyl ether, respectively) with structural similarities to triclosan. these strains showed less susceptibility to triclosan when grown in c ... | 2010 | 21041079 |
| structural insight into the expanded pcb-degrading abilities of a biphenyl dioxygenase obtained by directed evolution. | the biphenyl dioxygenase of burkholderia xenovorans lb400 is a multicomponent rieske-type oxygenase that catalyzes the dihydroxylation of biphenyl and many polychlorinated biphenyls (pcbs). the structural bases for the substrate specificity of the enzyme's oxygenase component (bphae(lb400)) are largely unknown. bphae(p4), a variant previously obtained through directed evolution, transforms several chlorobiphenyls, including 2,6-dichlorobiphenyl, more efficiently than bphae(lb400), yet differs fr ... | 2010 | 21073881 |
| anaerobic crystallization and initial x-ray diffraction data of biphenyl 2,3-dioxygenase from burkholderia xenovorans lb400: addition of agarose improved the quality of the crystals. | biphenyl 2,3-dioxygenase (bpdo; ec 1.14.12.18) catalyzes the initial step in the degradation of biphenyl and some polychlorinated biphenyls (pcbs). bpdolb400, the terminal dioxygenase component from burkholderia xenovorans lb400, a proteobacterial species that degrades a broad range of pcbs, has been crystallized under anaerobic conditions by sitting-drop vapour diffusion. initial crystals obtained using various polyethylene glycols as precipitating agents diffracted to very low resolution (∼8 å ... | 2010 | 21206025 |
| monitoring biodegradative enzymes with nanobodies raised in camelus dromedarius with mixtures of catabolic proteins. | functional studies of biodegradative activities in environmental microorganisms require molecular tools for monitoring catabolic enzymes in the members of the native microbiota. to this end, we have generated repertories of single-domain v(hh) fragments of camel immunoglobulins (nanobodies) able to interact with multiple proteins that are descriptors of environmentally relevant processes. for this, we immunized camelus dromedarius with a cocktail of up to 12 purified enzymes that are representat ... | 2011 | 21219561 |
| a product analog bound form of 3-oxoadipate-enol-lactonase (pcad) reveals a multifunctional role for the divergent cap domain. | lactones are a class of structurally diverse molecules that serve essential roles in biological processes ranging from quorum sensing to the aerobic catabolism of aromatic compounds. not surprisingly, enzymes involved in the bioprocessing of lactones are often targeted for protein engineering studies with the potential, for example, of optimized bioremediation of aromatic pollutants. the enol-lactone hydrolase (elh) represents one such class of targeted enzymes and catalyzes the conversion of 3- ... | 2011 | 21237173 |
| the homogentisate and homoprotocatechuate central pathways are involved in 3- and 4-hydroxyphenylacetate degradation by burkholderia xenovorans lb400. | genome characterization of the model pcb-degrading bacterium burkholderia xenovorans lb400 revealed the presence of eleven central pathways for aromatic compounds degradation, among them, the homogentisate and the homoprotocatechuate pathways. however, the functionality of these central pathways in strain lb400 has not been assessed and related peripheral pathways has not been described. | 2011 | 21423751 |
| elucidating the reaction mechanism of the benzoate oxidation pathway encoded aldehyde dehydrogenase from burkholderia xenovorans lb400. | oxidation of cis-3,4-dehydroadipyl-coa semialdehyde to cis-3,4-dehydroadipyl-coa by the aldehyde dehydrogenase, aldh(c) (ec.1.2.1.77), is an essential step in the metabolism of benzoate in burkholderia xenovorans lb400. in a previous study, we established a structural blueprint for this novel group of aldh enzymes. here, we build significantly on this initial work and propose a detailed reaction mechanism for aldh(c) based on comprehensive structural and functional investigations of active site ... | 2011 | 21495107 |
| genomic analysis of the phenylacetyl-coa pathway in burkholderia xenovorans lb400. | the phenylacetyl-coa (paa) catabolic pathway and genome-wide gene expression responses to phenylacetate catabolism were studied in the polychlorinated biphenyl (pcb)-degrading strain burkholderia xenovorans lb400. microarray and rt-qpcr analyses identified three non-contiguous chromosomal clusters of genes that are predicted to encode a complete paa pathway that were induced up to 40-fold during growth of lb400 on phenylacetate: paaghijkr, paaanebdf, and paac. comparison of the available genome ... | 2011 | 21519854 |
| retuning rieske-type oxygenases to expand substrate range. | rieske-type oxygenases are promising biocatalysts for the destruction of persistent pollutants or for the synthesis of fine chemicals. in this work, we explored pathways through which rieske-type oxygenases evolve to expand their substrate range. bphae(p4), a variant biphenyl dioxygenase generated from burkholderia xenovorans lb400 bphae(lb400) by the double substitution t335a/f336m, and bphae(rr41), obtained by changing asn(338), ile(341), and leu(409) of bphae(p4) to gln(338), val(341), and ph ... | 2011 | 21653696 |
| insight into the metabolism of 1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane (ddt) by biphenyl dioxygenases. | in this work we have investigated the ability of the biphenyl dioxygenase of burkholderia xenovorans lb400 (bphae(lb400)) and of pandoraea pnomenusa b356 (bphae(b356)) to metabolize ddt. data show bphae(lb400) is unable to metabolize this substrate but bphae(b356) metabolizes ddt to produce two stereoisomers. structural analysis of ddt-docked bphae(lb400) and bphae(b356) identified residue phe336 of bphae(lb400) as critical to prevent productive binding of ddt to bphae(lb400). furthermore, the f ... | 2011 | 22001737 |
| Isolation, characterization and docking studies of 2,3-dihydroxybiphenyl 1,2-dioxygenase from an activated sludge metagenome. | A 2,3-dihydroxybiphenyl-1,2-dioxygenase gene (designated as bphC_meta) was identified in activated sludge metagenome by PCR. This gene shared 99% sequence identity with BphC from Burkholderia xenovorans LB400. The enzyme was purified from recombinant Escherichia coli with a subunit molecular mass of 32 ± 1 kDa. It was optimally active at pH 9.0 and 40°C, using 2,3-dihydroxybiphenyl as a substrate. Activity toward substituted catechols was: 2,3-dihydroxybiphenyl > 3-methylcatechol > catechol > 4- ... | 2012 | 21898128 |
| adaptation mechanisms of bacteria during the degradation of polychlorinated biphenyls in the presence of natural and synthetic terpenes as potential degradation inducers. | in this study, we examined the effect of polychlorinated biphenyls (pcbs) in the presence of natural and synthetic terpenes and biphenyl on biomass production, lipid accumulation, and membrane adaptation mechanisms of two pcb-degrading bacterial strains pseudomonas stutzeri and burkholderia xenovorans lb400. according to the results obtained, it could be concluded that natural terpenes, mainly those contained in ivy leaves and pine needles, decreased adaptation responses induced by pcbs in these ... | 2011 | 22159613 |
| comparative genomics and functional analysis of the niap family uncover nicotinate transporters from bacteria, plants, and mammals. | the transporter(s) that mediate uptake of nicotinate and its n-methyl derivative trigonelline are not known in plants, and certain mammalian nicotinate transporters also remain unidentified. potential candidates for these missing transporters include proteins from the ubiquitous niap family. in bacteria, niap genes often belong to nad-related regulons, and genetic evidence supports a role for bacillus subtilis and acinetobacter baumannii niap proteins in uptake of nicotinate or nicotinamide. oth ... | 2011 | 21953179 |
| discovery of dna operators for tetr and marr family transcription factors fromburkholderia xenovorans. | determining transcription factor recognition motifs or operator sites is central to understanding gene regulation, yet few operators have been characterized. in this study, we used a protein-binding microarray (pbm) to discover the dna recognition sites and putative regulons for three tetr and one marr family tfs derived from b. xenovorans, which are common to the burkholderia genus. we also describe the development and application of a more streamlined version of the pbm technology that signifi ... | 2011 | 22117006 |
| Antioxidant compounds improved PCB-degradation by Burkholderia xenovorans strain LB400. | Polychlorobiphenyls (PCBs) are toxic and persistent organic pollutants that are widely distributed in the environment. Burkholderia xenovorans LB400 is capable of degrading aerobically an unusually wide range of PCBs. However, during PCB-degradation B. xenovorans LB400 generates reactive oxygen species (ROS) that affect its viability. The aim of this study was to increase the efficiency of PCB-degradation of B. xenovorans LB400 by adding antioxidant compounds that could increase tolerance to oxi ... | 2011 | 22142725 |