scholarly journals Mesaconase Activity of Class I Fumarase Contributes to Mesaconate Utilization by Burkholderia xenovorans

2015 ◽  
Vol 81 (16) ◽  
pp. 5632-5638 ◽  
Author(s):  
Miriam Kronen ◽  
Jahminy Sasikaran ◽  
Ivan A. Berg
Keyword(s):  
Energy Source ◽  
Pseudomonas Aeruginosa ◽  
Dicarboxylic Acid ◽  
Gene Cluster ◽  
Degradation Pathway ◽  
Class I ◽  
Acetyl Coa ◽  
Content Type ◽  
Burkholderia Xenovorans ◽  
Coa Transferase

ABSTRACTPseudomonas aeruginosa,Yersinia pestis, and many other bacteria are able to utilize the C5-dicarboxylic acid itaconate (methylenesuccinate). Itaconate degradation starts with its activation to itaconyl coenzyme A (itaconyl-CoA), which is further hydrated to (S)-citramalyl-CoA, and citramalyl-CoA is finally cleaved into acetyl-CoA and pyruvate. The xenobiotic-degrading betaproteobacteriumBurkholderia xenovoranspossesses aP. aeruginosa-like itaconate degradation gene cluster and is able to grow on itaconate and its isomer mesaconate (methylfumarate). Although itaconate degradation proceeds inB. xenovoransin the same way as inP. aeruginosa, the pathway of mesaconate utilization is not known. Here, we show that mesaconate is metabolized through its hydration to (S)-citramalate. The latter compound is then metabolized to acetyl-CoA and pyruvate with the participation of two enzymes of the itaconate degradation pathway, a promiscuous itaconate-CoA transferase able to activate (S)-citramalate in addition to itaconate and (S)-citramalyl-CoA lyase. The first reaction of the pathway, the mesaconate hydratase (mesaconase) reaction, is catalyzed by a class I fumarase. As this enzyme (Bxe_A3136) has similar efficiencies (kcat/Km) for both fumarate and mesaconate hydration, we conclude thatB. xenovoransclass I fumarase is in fact a promiscuous fumarase/mesaconase. This promiscuity is physiologically relevant, as it allows the growth of this bacterium on mesaconate as a sole carbon and energy source.

scholarly journals Steroid Degradation in Comamonas testosteroni TA441: Identification of the Entire β-Oxidation Cycle of the Cleaved B Ring

2019 ◽  
Vol 85 (20) ◽  
Author(s):  
Masae Horinouchi ◽  
Hiroyuki Koshino ◽  
Michal Malon ◽  
Hiroshi Hirota ◽  
Toshiaki Hayashi
Keyword(s):  
Gene Clusters ◽  
Degradation Process ◽  
Degradation Pathway ◽  
Ring Cleavage ◽  
Comamonas Testosteroni ◽  
Content Type ◽  
Coa Transferase ◽  
Degrading Bacteria ◽  
Globally Distributed

ABSTRACT Comamonas testosteroni TA441 degrades steroids via aromatization of the A ring, followed by degradation of 9,17-dioxo-1,2,3,4,10,19-hexanorandrostan-5-oic acid, mainly by β-oxidation. In this study, we revealed that 7β,9α-dihydroxy-17-oxo-1,2,3,4,10,19-hexanorandrostanoic acid-coenzyme A (CoA) ester is dehydrogenated by (3S)-3-hydroxylacyl CoA-dehydrogenase, encoded by scdE (ORF27), and then the resultant 9α-hydroxy-7,17-dioxo-1,2,3,4,10,19-hexanorandrostan-5-oic acid-CoA ester is converted by 3-ketoacyl-CoA transferase, encoded by scdF (ORF23). With these results, the whole cycle of β-oxidation on the side chain at C-8 of 9,17-dioxo-1,2,3,4,10,19-hexanorandrostan-5-oic acid is clarified; 9-hydroxy-17-oxo-1,2,3,4,10,19-hexanorandrostan-5-oic acid-CoA ester is dehydrogenated at C-6 by ScdC1C2, followed by hydration by ScdD. 7β,9α-Dihydroxy-17-oxo-1,2,3,4,10,19-hexanorandrostanoic acid-CoA ester then is dehydrogenated by ScdE to be converted to 9α-hydroxy-17-oxo-1,2,3,4,5,6,10,19-octanorandrostan-7-oic acid-CoA ester and acetyl-CoA by ScdF. ScdF is an ortholog of FadA6 in Mycobacterium tuberculosis H37Rv, which was reported as a 3-ketoacyl-CoA transferase involved in C ring cleavage. We also obtained results suggesting that ScdF is also involved in C ring cleavage, but further investigation is required for confirmation. ORF25 and ORF26, located between scdF and scdE, encode enzymes belonging to the amidase superfamily. Disrupting either ORF25 or ORF26 did not affect steroid degradation. Among the bacteria having gene clusters similar to those of tesB to tesR, some have both ORF25- and ORF26-like proteins or only an ORF26-like protein, but others do not have either ORF25- or ORF26-like proteins. ORF25 and ORF26 are not crucial for steroid degradation, yet they might provide clues to elucidate the evolution of bacterial steroid degradation clusters. IMPORTANCE Studies on bacterial steroid degradation were initiated more than 50 years ago primarily to obtain materials for steroid drugs. Steroid-degrading bacteria are globally distributed, and the role of bacterial steroid degradation in the environment as well as in relation to human health is attracting attention. The overall aerobic degradation of the four basic steroidal rings has been proposed; however, there is still much to be revealed to understand the complete degradation pathway. This study aims to uncover the whole steroid degradation process in Comamonas testosteroni TA441 as a model of steroid-degrading bacteria. C. testosteroni is one of the most studied representative steroid-degrading bacteria and is suitable for exploring the degradation pathway, because the involvement of degradation-related genes can be determined by gene disruption. Here, we elucidated the entire β-oxidation cycle of the cleaved B ring. This cycle is essential for the following C and D ring cleavage.

scholarly journals A Pathway for Degradation of Uracil to Acetyl Coenzyme A in Bacillus megaterium

2020 ◽  
Vol 86 (7) ◽  
Author(s):  
Di Zhu ◽  
Yifeng Wei ◽  
Jinyu Yin ◽  
Dazhi Liu ◽  
Ee Lui Ang ◽  
...  
Keyword(s):  
Energy Source ◽  
Bacillus Megaterium ◽  
Coenzyme A ◽  
Catabolic Pathway ◽  
Acetyl Coa ◽  
Acetyl Coenzyme A ◽  
Content Type ◽  
Biochemical Pathways ◽  
Acetyl Coenzyme

ABSTRACT Bacteria utilize diverse biochemical pathways for the degradation of the pyrimidine ring. The function of the pathways studied to date has been the release of nitrogen for assimilation. The most widespread of these pathways is the reductive pyrimidine catabolic pathway, which converts uracil into ammonia, carbon dioxide, and β-alanine. Here, we report the characterization of a β-alanine:pyruvate aminotransferase (PydD2) and an NAD+-dependent malonic semialdehyde dehydrogenase (MSDH) from a reductive pyrimidine catabolism gene cluster in Bacillus megaterium. Together, these enzymes convert β-alanine into acetyl coenzyme A (acetyl-CoA), a key intermediate in carbon and energy metabolism. We demonstrate the growth of B. megaterium in defined medium with uracil as its sole carbon and energy source. Homologs of PydD2 and MSDH are found in association with reductive pyrimidine pathway genes in many Gram-positive bacteria in the order Bacillales. Our study provides a basis for further investigations of the utilization of pyrimidines as a carbon and energy source by bacteria. IMPORTANCE Pyrimidine has wide occurrence in natural environments, where bacteria use it as a nitrogen and carbon source for growth. Detailed biochemical pathways have been investigated with focus mainly on nitrogen assimilation in the past decades. Here, we report the discovery and characterization of two important enzymes, PydD2 and MSDH, which constitute an extension for the reductive pyrimidine catabolic pathway. These two enzymes, prevalent in Bacillales based on our bioinformatics studies, allow stepwise conversion of β-alanine, a previous “end product” of the reductive pyrimidine degradation pathway, to acetyl-CoA as carbon and energy source.

scholarly journals Draft Genome Sequence of Pseudomonas aeruginosa Strain Hex1T Isolated from Soils Contaminated with Used Lubricating Oil in Argentina

2017 ◽  
Vol 5 (2) ◽  
Author(s):  
Adela M. Luján ◽  
Sofía Feliziani ◽  
Andrea M. Smania
Keyword(s):  
Energy Source ◽  
Pseudomonas Aeruginosa ◽  
Genome Sequence ◽  
Draft Genome ◽  
Heavy Metal Resistance ◽  
Metal Resistance ◽  
Draft Genome Sequence ◽  
Lubricating Oil ◽  
Content Type ◽  
Used Lubricating Oil

ABSTRACT Pseudomonas aeruginosa Hex1T was isolated from soils contaminated with used lubricating oil from a garage in Córdoba, Argentina. This strain is capable of utilizing this pollutant as the sole carbon and energy source. Here, we present the 6.9-Mb draft genome sequence of Hex1T, which contains many heavy metal-resistance genes.

scholarly journals The Hydroxyquinol Degradation Pathway in Rhodococcus jostii RHA1 and Agrobacterium Species Is an Alternative Pathway for Degradation of Protocatechuic Acid and Lignin Fragments

2020 ◽  
Vol 86 (19) ◽  
Author(s):  
Edward M. Spence ◽  
Heather T. Scott ◽  
Louison Dumond ◽  
Leonides Calvo-Bado ◽  
Sabrina di Monaco ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Protocatechuic Acid ◽  
Alternative Pathway ◽  
Degradation Pathway ◽  
Oxidative Decarboxylation ◽  
Content Type ◽  
Genes Encoding ◽  
Rhodococcus Jostii Rha1 ◽  
Similar Gene ◽  
Rhodococcus Jostii

ABSTRACT Deletion of the pcaHG genes, encoding protocatechuate 3,4-dioxygenase in Rhodococcus jostii RHA1, gives a gene deletion strain still able to grow on protocatechuic acid as the sole carbon source, indicating a second degradation pathway for protocatechuic acid. Metabolite analysis of wild-type R. jostii RHA1 grown on medium containing vanillin or protocatechuic acid indicated the formation of hydroxyquinol (benzene-1,2,4-triol) as a downstream product. Gene cluster ro01857-ro01860 in Rhodococcus jostii RHA1 contains genes encoding hydroxyquinol 1,2-dioxygenase and maleylacetate reductase for degradation of hydroxyquinol but also putative mono-oxygenase (ro01860) and putative decarboxylase (ro01859) genes, and a similar gene cluster is found in the genome of lignin-degrading Agrobacterium species. Recombinant R. jostii mono-oxygenase and decarboxylase enzymes in combination were found to convert protocatechuic acid to hydroxyquinol. Hence, an alternative pathway for degradation of protocatechuic acid via oxidative decarboxylation to hydroxyquinol is proposed. IMPORTANCE There is a well-established paradigm for degradation of protocatechuic acid via the β-ketoadipate pathway in a range of soil bacteria. In this study, we have found the existence of a second pathway for degradation of protocatechuic acid in Rhodococcus jostii RHA1, via hydroxyquinol (benzene-1,2,4-triol), which establishes a metabolic link between protocatechuic acid and hydroxyquinol. The presence of this pathway in a lignin-degrading Agrobacterium sp. strain suggests the involvement of the hydroxyquinol pathway in the metabolism of degraded lignin fragments.

The pimFABCDE operon from Rhodopseudomonas palustris mediates dicarboxylic acid degradation and participates in anaerobic benzoate degradation

Microbiology ◽  
2005 ◽  
Vol 151 (3) ◽  
pp. 727-736 ◽  
Author(s):  
Faith H. Harrison ◽  
Caroline S. Harwood
Keyword(s):  
Fatty Acids ◽  
Dicarboxylic Acid ◽  
Gene Cluster ◽  
Rhodopseudomonas Palustris ◽  
Dicarboxylic Acids ◽  
Wild Type ◽  
Medium Chain Length ◽  
Coa Ligase ◽  
Anoxic Environments ◽  
Coa Transferase

Bacteria in anoxic environments typically convert aromatic compounds derived from pollutants or green plants to benzoyl-CoA, and then to the C7 dicarboxylic acid derivative 3-hydroxypimelyl-CoA. Inspection of the recently completed genome sequence of the purple nonsulfur phototroph Rhodopseudomonas palustris revealed one predicted cluster of genes for the β-oxidation of dicarboxylic acids. These genes, annotated as pimFABCDE, are predicted to encode acyl-CoA ligase, enoyl-CoA hydratase, acyl-CoA dehydrogenase and acyl-CoA transferase enzymes, which should allow the conversion of odd-chain dicarboxylic acids to glutaryl-CoA, and even-chain dicarboxylic acids to succinyl-CoA. A mutant strain that was deleted in the pim gene cluster grew at about half the rate of the wild-type parent when benzoate or pimelate was supplied as the sole carbon source. The mutant grew five times more slowly than the wild-type on the C14 dicarboxylic acid tetradecanedioate. The mutant was unimpaired in growth on the C8-fatty acid caprylate. The acyl-CoA ligase predicted to be encoded by the pimA gene was purified, and found to be active with C7–C14 dicarboxylic and fatty acids. The expression of a pimA–lacZ chromosomal gene fusion increased twofold when cells were grown in the presence of straight-chain C7–C14 dicarboxylic and fatty acids. These results suggest that the β-oxidation enzymes encoded by the pim gene cluster are active with medium-chain-length dicarboxylic acids, including pimelate. However, the finding that the pim operon deletion mutant is still able to grow on dicarboxylic acids, albeit at a slower rate, indicates that R. palustris has additional genes that can also specify the degradation of these compounds.

scholarly journals Ethylene Glycol Metabolism in the Acetogen Acetobacterium woodii

2016 ◽  
Vol 198 (7) ◽  
pp. 1058-1065 ◽  
Author(s):  
Dragan Trifunović ◽  
Kai Schuchmann ◽  
Volker Müller
Keyword(s):  
Ethylene Glycol ◽  
Gene Cluster ◽  
Dual Function ◽  
Acetobacterium Woodii ◽  
Acetyl Coa ◽  
Terminal Electron Acceptor ◽  
Content Type ◽  
Bacterial Microcompartments ◽  
Genes Encoding ◽  
Acetogenic Bacterium

ABSTRACTThe acetogenic bacteriumAcetobacterium woodiiis able to grow by the oxidation of diols, such as 1,2-propanediol, 2,3-butanediol, or ethylene glycol. Recent analyses demonstrated fundamentally different ways for oxidation of 1,2-propanediol and 2,3-butanediol. Here, we analyzed the metabolism of ethylene glycol. Our data demonstrate that ethylene glycol is dehydrated to acetaldehyde, which is then disproportionated to ethanol and acetyl coenzyme A (acetyl-CoA). The latter is further converted to acetate, and this pathway is coupled to ATP formation by substrate-level phosphorylation. Apparently, the product ethanol is in part further oxidized and the reducing equivalents are recycled by reduction of CO2to acetate in the Wood-Ljungdahl pathway. Biochemical data as well as the results of protein synthesis analysis are consistent with the hypothesis that the propane diol dehydratase (PduCDE) and CoA-dependent propionaldehyde dehydrogenase (PduP) proteins, encoded by thepdugene cluster, also catalyze ethylene glycol dehydration to acetaldehyde and its CoA-dependent oxidation to acetyl-CoA. Moreover, genes encoding bacterial microcompartments as part of thepdugene cluster are also expressed during growth on ethylene glycol, arguing for a dual function of the Pdu microcompartment system.IMPORTANCEAcetogenic bacteria are characterized by their ability to use CO2as a terminal electron acceptor by a specific pathway, the Wood-Ljungdahl pathway, enabling in most acetogens chemolithoautotrophic growth with H2and CO2. However, acetogens are very versatile and can use a wide variety of different substrates for growth. Here we report on the elucidation of the pathway for utilization of ethylene glycol by the model acetogenAcetobacterium woodii. This diol is degraded by dehydration to acetaldehyde followed by a disproportionation to acetate and ethanol. We present evidence that this pathway is catalyzed by the same enzyme system recently described for the utilization of 1,2-propanediol. The enzymes for ethylene glycol utilization seem to be encapsulated in protein compartments, known as bacterial microcompartments.

scholarly journals Discovery and Biosynthesis of the Antibiotic Bicyclomycin in Distantly Related Bacterial Classes

2018 ◽  
Vol 84 (9) ◽  
Author(s):  
Natalia M. Vior ◽  
Rodney Lacret ◽  
Govind Chandra ◽  
Siobhán Dorai-Raj ◽  
Martin Trick ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Gene Cluster ◽  
Gene Clusters ◽  
Opportunistic Pathogen ◽  
Biosynthetic Gene Cluster ◽  
Wide Distribution ◽  
Biosynthetic Gene ◽  
P450 Monooxygenase ◽  
Content Type ◽  
Transcription Termination Factor

ABSTRACTBicyclomycin (BCM) is a clinically promising antibiotic that is biosynthesized byStreptomyces cinnamoneusDSM 41675. BCM is structurally characterized by a core cyclo(l-Ile-l-Leu) 2,5-diketopiperazine (DKP) that is extensively oxidized. Here, we identify the BCM biosynthetic gene cluster, which shows that the core of BCM is biosynthesized by a cyclodipeptide synthase, and the oxidative modifications are introduced by five 2-oxoglutarate-dependent dioxygenases and one cytochrome P450 monooxygenase. The discovery of the gene cluster enabled the identification of BCM pathways encoded by the genomes of hundreds ofPseudomonas aeruginosaisolates distributed globally, and heterologous expression of the pathway fromP. aeruginosaSCV20265 demonstrated that the product is chemically identical to BCM produced byS. cinnamoneus. Overall, putative BCM gene clusters have been found in at least seven genera spanningActinobacteriaandProteobacteria(Alphaproteobacteria,Betaproteobacteria, andGammaproteobacteria). This represents a rare example of horizontal gene transfer of an intact biosynthetic gene cluster across such distantly related bacteria, and we show that these gene clusters are almost always associated with mobile genetic elements.IMPORTANCEBicyclomycin is the only natural product antibiotic that selectively inhibits the transcription termination factor Rho. This mechanism of action, combined with its proven biological safety and its activity against clinically relevant Gram-negative bacterial pathogens, makes it a very promising antibiotic candidate. Here, we report the identification of the bicyclomycin biosynthetic gene cluster in the known bicyclomycin-producing organismStreptomyces cinnamoneus, which will enable the engineered production of new bicyclomycin derivatives. The identification of this gene cluster also led to the discovery of hundreds of bicyclomycin pathways encoded in highly diverse bacteria, including in the opportunistic pathogenPseudomonas aeruginosa. This wide distribution of a complex biosynthetic pathway is very unusual and provides an insight into how a pathway for an antibiotic can be transferred between diverse bacteria.

scholarly journals DbdR, a New Member of the LysR Family of Transcriptional Regulators, Coordinately Controls Four Promoters in theThauera aromaticaAR-1 3,5-Dihydroxybenzoate Anaerobic Degradation Pathway

2018 ◽  
Vol 85 (2) ◽  
Author(s):  
Daniel Pacheco-Sánchez ◽  
Águeda Molina-Fuentes ◽  
Patricia Marín ◽  
Alberto Díaz-Romero ◽  
Silvia Marqués
Keyword(s):  
Energy Source ◽  
Binding Site ◽  
Anaerobic Degradation ◽  
Degradation Pathway ◽  
Anaerobic Growth ◽  
Nitrate Respiration ◽  
Facultative Anaerobe ◽  
Content Type ◽  
Basal Expression ◽  
Thauera Aromatica

ABSTRACTThe facultative anaerobeThauera aromaticastrain AR-1 uses 3,5-dihydroxybenzoate (3,5-DHB) as a sole carbon and energy source under anoxic conditions using an unusual oxidative strategy to overcome aromatic ring stability. A 25-kb gene cluster organized in four main operons encodes the anaerobic degradation pathway for this aromatic. ThedbdRgene coding for a LysR-type transcriptional regulator (LTTR), which is present at the foremost end of the cluster, is required for anaerobic growth on 3,5-DHB and for the expression of the main pathway operons. A model structure of DbdR showed conserved key residues for effector binding with its closest relative TsaR forp-toluenesulfonate degradation. We found that DbdR controlled expression of three promoters upstream from the operons coding for the three main steps of the pathway. While one of them (Porf20) was only active in the presence of 3,5-DHB, the other two (PdbhLand Porf18) showed moderate basal levels that were further induced in the presence of the pathway substrate, which needed be converted to hydroxyhydroquinone to activate transcription. Both basal and induced activities were strictly dependent on DbdR, which was also required for transcription from its own promoter. DbdR basal expression was moderately high and, unlike most LTTR, increased 2-fold in response to the presence of the effector. DbdR was found to be a tetramer in solution, producing a single retardation complex in binding assays with the three enzymatic promoters, consistent with its tetrameric structure. The three promoters had a conserved organization with a clear putative primary (regulatory) binding site and a putative secondary (activating) binding site positioned at the expected distances from the transcription start site. In contrast, two protein-DNA complexes were observed for the PdbdRpromoter, which also showed significant sequence divergence from those of the three other promoters. Taken together, our results show that a single LTTR coordinately controls expression of the entire 3,5-DHB anaerobic degradation pathway inThauera aromaticaAR-1, allowing a fast and optimized response to the presence of the aromatic.IMPORTANCEThauera aromaticaAR-1 is a facultative anaerobe that is able to use 3,5-dihydroxybenzoat (3,5-DHB) as the sole carbon and energy source in a process that is dependent on nitrate respiration. We have shown that a single LysR-type regulator with unusual properties, DbdR, controls the expression of the pathway in response to the presence of the substrate; unlike other regulators of the family, DbdR does not repress but activates its own synthesis and is able to bind and activate three promoters directing the synthesis of the pathway enzymes. The promoter architecture is conserved among the three promoters but deviates from that of typical LTTR-dependent promoters. The substrate must be metabolized to an intermediate compound to activate transcription, which requires basal enzyme levels to always be present. The regulatory network present in this strain is designed to allow basal expression of the enzymatic machinery, which would rapidly metabolize the substrate when exposed to it, thus rendering the effector molecule. Once activated, the regulator induces the synthesis of the entire pathway through a positive feedback, increasing expression from all the target promoters to allow maximum growth.

scholarly journals Catabolism of 2-Hydroxypyridine byBurkholderiasp. Strain MAK1: a 2-Hydroxypyridine 5-Monooxygenase Encoded byhpdABCDECatalyzes the First Step of Biodegradation

2018 ◽  
Vol 84 (11) ◽  
Author(s):  
Vytautas Petkevičius ◽  
Justas Vaitekūnas ◽  
Jonita Stankevičiūtė ◽  
Renata Gasparavičiūtė ◽  
Rolandas Meškys
Keyword(s):  
Gene Cluster ◽  
Heterocyclic Compounds ◽  
Degradation Pathway ◽  
Blue Pigment ◽  
Pyridine Derivatives ◽  
Catabolic Pathway ◽  
Content Type ◽  
Key Enzyme ◽  
Regioselective Hydroxylation ◽  
Dna Fragment

ABSTRACTMicrobial degradation of 2-hydroxypyridine usually results in the formation of a blue pigment (nicotine blue). In contrast, theBurkholderiasp. strain MAK1 bacterium utilizes 2-hydroxypyridine without the accumulation of nicotine blue. This scarcely investigated degradation pathway presumably employs 2-hydroxypyridine 5-monooxygenase, an elusive enzyme that has been hypothesized but has yet to be identified or characterized. The isolation of the mutant strainBurkholderiasp. MAK1 ΔP5 that is unable to utilize 2-hydroxypyridine has led to the identification of a gene cluster (designatedhpd) which is responsible for the degradation of 2-hydroxypyridine. The activity of 2-hydroxypyridine 5-monooxygenase has been assigned to a soluble diiron monooxygenase (SDIMO) encoded by a five-gene cluster (hpdA,hpdB,hpdC,hpdD, andhpdE). A 4.5-kb DNA fragment containing all five genes has been successfully expressed inBurkholderiasp. MAK1 ΔP5 cells. We have proved that the recombinant HpdABCDE protein catalyzes the enzymatic turnover of 2-hydroxypyridine to 2,5-dihydroxypyridine. Moreover, we have confirmed that emerging 2,5-dihydroxypyridine is a substrate for HpdF, an enzyme similar to 2,5-dihydroxypyridine 5,6-dioxygenases that are involved in the catabolic pathways of nicotine and nicotinic acid. The proteins and genes identified in this study have allowed the identification of a novel degradation pathway of 2-hydroxypyridine. Our results provide a better understanding of the biodegradation of pyridine derivatives in nature. Also, the discovered 2-hydroxypyridine 5-monooxygenase may be an attractive catalyst for the regioselective synthesis of variousN-heterocyclic compounds.IMPORTANCEThe degradation pathway of 2-hydroxypyridine without the accumulation of a blue pigment is relatively unexplored, as, to our knowledge, no genetic data related to this process have ever been presented. In this paper, we describe genes and enzymes involved in this little-studied catabolic pathway. This work provides new insights into the metabolism of 2-hydroxypyridine in nature. A broad-range substrate specificity of 2-hydroxypyridine 5-monooxygenase, a key enzyme in the degradation, makes this biocatalyst attractive for the regioselective hydroxylation of pyridine derivatives.

scholarly journals Five New Genes Are Important for Common Polysaccharide Antigen Biosynthesis in Pseudomonas aeruginosa

mBio ◽  
2013 ◽  
Vol 4 (1) ◽  
Author(s):  
Youai Hao ◽  
Jerry D. King ◽  
Steven Huszczynski ◽  
Dana Kocíncová ◽  
Joseph S. Lam
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Cell Surface ◽  
Gene Cluster ◽  
Opportunistic Pathogen ◽  
Airway Epithelial Cells ◽  
Airway Epithelial ◽  
Polysaccharide Antigen ◽  
Content Type ◽  
New Genes ◽  
Cell Surface Structure

ABSTRACTCommon polysaccharide antigen (CPA) is a conserved cell surface polysaccharide produced byPseudomonas aeruginosa. It contains a rhamnan homopolymer and is one of the two forms of O polysaccharide attached toP. aeruginosalipopolysaccharide (LPS). Our laboratory has previously characterized an eight-gene cluster (pa5447-pa5454inP. aeruginosaPAO1) required for biosynthesis of CPA. Here we demonstrate that an adjacent five-gene clusterpa5455-pa5459is also involved. Using reverse transcriptase PCR (RT-PCR), we showed that the original eight-gene cluster and the new five-gene cluster are both organized as operons. We have analyzed the LPS phenotypes of in-frame deletion mutants made in each of the five genes, and the results verified that these five genes are indeed required for CPA biosynthesis, extending the CPA biosynthesis locus to contain 13 contiguous genes. By performing overexpression experiments of different sets of these biosynthesis genes, we were able to obtain information about their possible functions in CPA biosynthesis.IMPORTANCELipopolysaccharide (LPS) is an important cell surface structure of Gram-negative bacteria. The human opportunistic pathogenPseudomonas aeruginosasimultaneously produces an O-antigen-specific (OSA) form and a common polysaccharide antigen (CPA) form of LPS. CPA, the focus of this study, is composed of α-1-2, α1-3-linkedd-rhamnose sugars and has been shown to be important for attachment of the bacteria to human airway epithelial cells. Genome sequencing of this species revealed a new five-gene cluster that we predicted to be involved in CPA biosynthesis and modification. In this study, we have generated chromosomal knockouts by performing in-frame deletions and allelic replacements. Characterizing the function of each of the five genes is important for us to better understand CPA biosynthesis and the mechanisms of chain length termination and regulation of this unique D-rhamnan polysaccharide.

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