Degradation of Benzene by Pseudomonas veronii 1YdBTEX2 and 1YB2 Is Catalyzed by Enzymes Encoded in Distinct Catabolism Gene Clusters

ABSTRACTPseudomonas veronii1YdBTEX2, a benzene and toluene degrader, andPseudomonas veronii1YB2, a benzene degrader, have previously been shown to be key players in a benzene-contaminated site. These strains harbor unique catabolic pathways for the degradation of benzene comprising a gene cluster encoding an isopropylbenzene dioxygenase where genes encoding downstream enzymes were interrupted by stop codons. Extradiol dioxygenases were recruited from gene clusters comprising genes encoding a 2-hydroxymuconic semialdehyde dehydrogenase necessary for benzene degradation but typically absent from isopropylbenzene dioxygenase-encoding gene clusters. The benzene dihydrodiol dehydrogenase-encoding gene was not clustered with any other aromatic degradation genes, and the encoded protein was only distantly related to dehydrogenases of aromatic degradation pathways. The involvement of the different gene clusters in the degradation pathways was suggested by real-time quantitative reverse transcription PCR.

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Two Gene Clusters Coordinate Galactose and Lactose Metabolism in Streptococcus gordonii

Applied and Environmental Microbiology ◽

10.1128/aem.01393-12 ◽

2012 ◽

Vol 78 (16) ◽

pp. 5597-5605 ◽

Cited By ~ 28

Author(s):

Lin Zeng ◽

Nicole C. Martino ◽

Robert A. Burne

Keyword(s):

Gene Clusters ◽

Selective Advantage ◽

Streptococcus Gordonii ◽

Phosphotransferase System ◽

Content Type ◽

High Affinity ◽

Oral Biofilms ◽

Genes Encoding ◽

Complex Regulation ◽

Galactose Utilization

ABSTRACTStreptococcus gordoniiis an early colonizer of the human oral cavity and an abundant constituent of oral biofilms. Two tandemly arranged gene clusters, designatedlacandgal, were identified in theS. gordoniiDL1 genome, which encode genes of the tagatose pathway (lacABCD) and sugar phosphotransferase system (PTS) enzyme II permeases. Genes encoding a predicted phospho-β-galactosidase (LacG), a DeoR family transcriptional regulator (LacR), and a transcriptional antiterminator (LacT) were also present in the clusters. Growth and PTS assays supported that the permease designated EIILactransports lactose and galactose, whereas EIIGaltransports galactose. The expression of the gene for EIIGalwas markedly upregulated in cells growing on galactose. Using promoter-catfusions, a role for LacR in the regulation of the expressions of both gene clusters was demonstrated, and thegalcluster was also shown to be sensitive to repression by CcpA. The deletion oflacTcaused an inability to grow on lactose, apparently because of its role in the regulation of the expression of the genes for EIILac, but had little effect on galactose utilization.S. gordoniimaintained a selective advantage overStreptococcus mutansin a mixed-species competition assay, associated with its possession of a high-affinity galactose PTS, althoughS. mutanscould persist better at low pHs. Collectively, these results support the concept that the galactose and lactose systems ofS. gordoniiare subject to complex regulation and that a high-affinity galactose PTS may be advantageous whenS. gordoniiis competing against the caries pathogenS. mutansin oral biofilms.

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Redundancy in Aromatic O-Demethylation and Ring-Opening Reactions in Novosphingobium aromaticivorans and Their Impact in the Metabolism of Plant-Derived Phenolics

Applied and Environmental Microbiology ◽

10.1128/aem.02794-20 ◽

2021 ◽

Vol 87 (8) ◽

Author(s):

Jose M. Perez ◽

Wayne S. Kontur ◽

Carson Gehl ◽

Derek M. Gille ◽

Yanjun Ma ◽

...

Keyword(s):

Phenolic Compounds ◽

Substrate Specificity ◽

Aromatic Ring ◽

Ring Opening ◽

Plant Biomass ◽

Syringic Acid ◽

Detailed Knowledge ◽

Degradation Pathways ◽

Content Type ◽

Aromatic Degradation

ABSTRACT Lignin is a plant heteropolymer composed of phenolic subunits. Because of its heterogeneity and recalcitrance, the development of efficient methods for its valorization still remains an open challenge. One approach to utilize lignin is its chemical deconstruction into mixtures of monomeric phenolic compounds, followed by biological funneling into a single product. Novosphingobium aromaticivorans DSM 12444 has been previously engineered to produce 2-pyrone-4,6-dicarboxylic acid (PDC) from depolymerized lignin by simultaneously metabolizing multiple aromatics through convergent routes involving the intermediates 3-methoxygallic acid (3-MGA) and protocatechuic acid (PCA). We investigated enzymes predicted to be responsible for O-demethylation and oxidative aromatic ring opening, two critical reactions involved in the metabolism of phenolic compounds by N. aromaticivorans. The results showed the involvement of DesA in O-demethylation of syringic and vanillic acids, LigM in O-demethylation of vanillic acid and 3-MGA, and a new O-demethylase, DmtS, in the conversion of 3-MGA into gallic acid (GA). In addition, we found that LigAB was the main aromatic ring-opening dioxygenase involved in 3-MGA, PCA, and GA metabolism and that a previously uncharacterized dioxygenase, LigAB2, had high activity with GA. Our results indicate a metabolic route not previously identified in N. aromaticivorans that involves O-demethylation of 3-MGA to GA. We predict that this pathway channels ∼15% of the carbon flow from syringic acid, with the rest following ring opening of 3-MGA. The new knowledge obtained in this study allowed for the creation of an improved engineered strain for the funneling of aromatic compounds that exhibits stoichiometric conversion of syringic acid into PDC. IMPORTANCE For lignocellulosic biorefineries to effectively contribute to reduction of fossil fuel use, they need to become efficient at producing chemicals from all major components of plant biomass. Making products from lignin will require engineering microorganisms to funnel multiple phenolic compounds to the chemicals of interest, and N. aromaticivorans is a promising chassis for this technology. The ability of N. aromaticivorans to efficiently and simultaneously degrade many phenolic compounds may be linked to having functionally redundant aromatic degradation pathways and enzymes with broad substrate specificity. A detailed knowledge of aromatic degradation pathways is thus essential to identify genetic engineering targets to maximize product yields. Furthermore, knowledge of enzyme substrate specificity is critical to redirect flow of carbon to desired pathways. This study described an uncharacterized pathway in N. aromaticivorans and the enzymes that participate in this pathway, allowing the engineering of an improved strain for production of PDC from lignin.

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Employing a Recombinant Strain of Advenella mimigardefordensis for Biotechnical Production of Homopolythioesters from 3,3′-Dithiodipropionic Acid

Applied and Environmental Microbiology ◽

10.1128/aem.00007-12 ◽

2012 ◽

Vol 78 (9) ◽

pp. 3286-3297 ◽

Cited By ~ 17

Author(s):

Yongzhen Xia ◽

Jan Hendrik Wübbeler ◽

Qingsheng Qi ◽

Alexander Steinbüchel

Keyword(s):

Carbon Source ◽

Dry Weight ◽

Pha Synthase ◽

Mineral Salts ◽

Content Type ◽

Genes Encoding ◽

Polyhydroxyalkanoate Synthase ◽

Encoding Gene ◽

Different Origins ◽

Severe Growth

ABSTRACTAdvenella mimigardefordensisstrain DPN7Twas genetically modified to produce poly(3-mercaptopropionic acid) (PMP) homopolymer by exploiting the recently unraveled process of 3,3′-dithiodipropionic acid (DTDP) catabolism. Production was achieved by systematically engineering the metabolism of this strain as follows: (i) deletion of its inherent 3MP dioxygenase-encoding gene (mdo), (ii) introduction of thebuk-ptboperon (genes encoding the butyrate kinase, Buk, and the phosphotransbutyrylase, Ptb, fromClostridium acetobutylicum), and (iii) overexpression of its own polyhydroxyalkanoate synthase (phaCAm). These measures yielded the potent PMP production strainA. mimigardefordensisstrain SHX22. The deletion ofmdowas required for adequate synthesis of PMP due to the resulting accumulation of 3MP during utilization of DTDP. Overexpression of the plasmid-bornebuk-ptboperon caused a severe growth repression. This effect was overcome by inserting this operon into the genome. Polyhydroxyalkanoate (PHA) synthases from different origins were compared. The native PHA synthase ofA. mimigardefordensis(phaCAm) was obviously the best choice to establish homopolythioester production in this strain. In addition, the cultivation conditions, including an appropriate provision of the carbon source, were further optimized to enhance PMP production. The engineered strain accumulated PMP up to approximately 25% (wt/wt) of the cell dry weight when cultivated in mineral salts medium containing glycerol as the carbon source in addition to DTDP as the sulfur-providing precursor. According to our knowledge, this is the first report of PMP homopolymer production by a metabolically engineered bacterium using DTDP, which is nontoxic, as the precursor substrate.

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Glycosulfatase-Encoding Gene Cluster in Bifidobacterium breve UCC2003

Applied and Environmental Microbiology ◽

10.1128/aem.02022-16 ◽

2016 ◽

Vol 82 (22) ◽

pp. 6611-6623 ◽

Cited By ~ 23

Author(s):

Muireann Egan ◽

Hao Jiang ◽

Mary O'Connell Motherway ◽

Stefan Oscarson ◽

Douwe van Sinderen

Keyword(s):

Gastrointestinal Tract ◽

Gut Microbiota ◽

Gene Cluster ◽

Digestive Tract ◽

Gene Clusters ◽

Content Type ◽

Reading Frame ◽

Bifidobacterium Breve ◽

Breastfed Infants ◽

Encoding Gene

ABSTRACTBifidobacteria constitute a specific group of commensal bacteria typically found in the gastrointestinal tract (GIT) of humans and other mammals.Bifidobacterium brevestrains are numerically prevalent among the gut microbiota of many healthy breastfed infants. In the present study, we investigated glycosulfatase activity in a bacterial isolate from a nursling stool sample,B. breveUCC2003. Two putative sulfatases were identified on the genome ofB. breveUCC2003. The sulfated monosaccharideN-acetylglucosamine-6-sulfate (GlcNAc-6-S) was shown to support the growth ofB. breveUCC2003, whileN-acetylglucosamine-3-sulfate,N-acetylgalactosamine-3-sulfate, andN-acetylgalactosamine-6-sulfate did not support appreciable growth. By using a combination of transcriptomic and functional genomic approaches, a gene cluster designatedats2was shown to be specifically required for GlcNAc-6-S metabolism. Transcription of theats2cluster is regulated by a repressor open reading frame kinase (ROK) family transcriptional repressor. This study represents the first description of glycosulfatase activity within theBifidobacteriumgenus.IMPORTANCEBifidobacteria are saccharolytic organisms naturally found in the digestive tract of mammals and insects.Bifidobacterium brevestrains utilize a variety of plant- and host-derived carbohydrates that allow them to be present as prominent members of the infant gut microbiota as well as being present in the gastrointestinal tract of adults. In this study, we introduce a previously unexplored area of carbohydrate metabolism in bifidobacteria, namely, the metabolism of sulfated carbohydrates.B. breveUCC2003 was shown to metabolizeN-acetylglucosamine-6-sulfate (GlcNAc-6-S) through one of two sulfatase-encoding gene clusters identified on its genome. GlcNAc-6-S can be found in terminal or branched positions of mucin oligosaccharides, the glycoprotein component of the mucous layer that covers the digestive tract. The results of this study provide further evidence of the ability of this species to utilize mucin-derived sugars, a trait which may provide a competitive advantage in both the infant gut and adult gut.

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Off-Target Gene Regulation Mediated by Transcriptional Repressors of Antimicrobial Efflux Pump Genes in Neisseria gonorrhoeae

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00010-11 ◽

2011 ◽

Vol 55 (6) ◽

pp. 2559-2565 ◽

Cited By ~ 12

Author(s):

Paul J. T. Johnson ◽

Virginia A. Stringer ◽

William M. Shafer

Keyword(s):

Dna Binding ◽

Neisseria Gonorrhoeae ◽

Binding Proteins ◽

Efflux Pump ◽

Dna Binding Proteins ◽

Transcriptional Repressors ◽

Content Type ◽

Genes Encoding ◽

Encoding Gene ◽

Microarray Studies

ABSTRACTDNA-binding proteins that control expression of drug efflux pump genes have been termed “local regulators” as their encoding gene is often located adjacent to the gene(s) that they regulate. However, results from recent studies indicate that they can control genes outside efflux pump-encoding loci, which we term as being “off target.” For example, the MtrR repressor was initially recognized for its ability to repress transcription of themtrCDE-encoded efflux pump operon in the strict human pathogenNeisseria gonorrhoeae, but recent results from genetic and microarray studies have shown that it can control expression of nearly 70 genes scattered throughout the chromosome. One of the off-target MtrR-repressed genes isglnA, which encodes glutamine synthetase. Herein, we confirm the capacity of MtrR to repressglnAexpression and provide evidence that such repression is due to its ability to negatively influence the binding of a second DNA-binding protein (FarR), which activatesglnA. FarR was previously recognized as a transcriptional repressor of thefarAB-encoded efflux pump operon. Thus, two DNA-binding proteins previously characterized as repressors of genes encoding efflux pumps that contribute to gonococcal resistance to antimicrobials can act in an opposing manner to modulate expression of a gene involved in basic metabolism.

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Draft Genome Sequence of Phenol-Degrading Variovorax boronicumulans Strain HAB-30

Microbiology Resource Announcements ◽

10.1128/mra.01478-19 ◽

2020 ◽

Vol 9 (7) ◽

Author(s):

Fatma Azwani Abdul Aziz ◽

Kenshi Suzuki ◽

Ryota Moriuchi ◽

Hideo Dohra ◽

Yosuke Tashiro ◽

...

Keyword(s):

Genome Sequence ◽

Aromatic Compounds ◽

Metabolic Pathways ◽

Chemostat Culture ◽

Phenol Degradation ◽

Draft Genome ◽

Draft Genome Sequence ◽

Degradation Pathways ◽

Content Type ◽

Genes Encoding

We report the draft genome sequence of Variovorax boronicumulans strain HAB-30, which was isolated from a phenol-degrading chemostat culture. This strain contains genes encoding a multicomponent type of phenol hydroxylase, with degradation pathways for catechol and other aromatic compounds. The genome sequence will be useful for understanding the metabolic pathways involved in phenol degradation.

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Genetic Analysis Reveals the Identity of the Photoreceptor for Phototaxis in Hormogonium Filaments of Nostoc punctiforme

Journal of Bacteriology ◽

10.1128/jb.02374-14 ◽

2014 ◽

Vol 197 (4) ◽

pp. 782-791 ◽

Cited By ~ 35

Author(s):

Elsie L. Campbell ◽

Kari D. Hagen ◽

Rui Chen ◽

Douglas D. Risser ◽

Daniela P. Ferreira ◽

...

Keyword(s):

Mutational Analysis ◽

Transcriptional Start Site ◽

Gene Clusters ◽

Gliding Motility ◽

Transcriptional Analysis ◽

Nostoc Punctiforme ◽

Recognition Sequence ◽

Content Type ◽

Wide Range ◽

Genes Encoding

In cyanobacterialNostocspecies, substratum-dependent gliding motility is confined to specialized nongrowing filaments called hormogonia, which differentiate from vegetative filaments as part of a conditional life cycle and function as dispersal units. Here we confirm thatNostoc punctiformehormogonia are positively phototactic to white light over a wide range of intensities.N. punctiformecontains two gene clusters (clusters 2 and 2i), each of which encodes modular cyanobacteriochrome–methyl-accepting chemotaxis proteins (MCPs) and other proteins that putatively constitute a basic chemotaxis-like signal transduction complex. Transcriptional analysis established that all genes in clusters 2 and 2i, plus two additional clusters (clusters 1 and 3) with genes encoding MCPs lacking cyanobacteriochrome sensory domains, are upregulated during the differentiation of hormogonia. Mutational analysis determined that only genes in cluster 2i are essential for positive phototaxis inN. punctiformehormogonia; here these genes are designatedptx(forphototaxis) genes. The cluster is unusual in containing complete or partial duplicates of genes encoding proteins homologous to the well-described chemotaxis elements CheY, CheW, MCP, and CheA. The cyanobacteriochrome-MCP gene (ptxD) lacks transmembrane domains and has 7 potential binding sites for bilins. The transcriptional start site of theptxgenes does not resemble a sigma 70 consensus recognition sequence; moreover, it is upstream of two genes encoding gas vesicle proteins (gvpAandgvpC), which also are expressed only in the hormogonium filaments ofN. punctiforme.

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Polycyclic Aromatic Hydrocarbon (PAH) Degradation Pathways of the Obligate Marine PAH DegraderCycloclasticussp. Strain P1

Applied and Environmental Microbiology ◽

10.1128/aem.01261-18 ◽

2018 ◽

Vol 84 (21) ◽

Cited By ~ 10

Author(s):

Wanpeng Wang ◽

Lin Wang ◽

Zongze Shao

Keyword(s):

Metabolic Network ◽

Aromatic Hydrocarbons ◽

Gene Clusters ◽

Degradation Pathway ◽

Degradation Pathways ◽

Marine Environments ◽

Content Type ◽

Pah Degradation ◽

Polycyclic Aromatic ◽

Pah Metabolism

ABSTRACTBacteria play an important role in the removal of polycyclic aromatic hydrocarbons (PAHs) from polluted environments. In marine environments,Cycloclasticusis one of the most prevalent PAH-degrading bacterial genera. However, little is known regarding the degradation mechanisms for multiple PAHs byCycloclasticus.Cycloclasticussp. strain P1 was isolated from deep-sea sediments and is known to degrade naphthalene, phenanthrene, pyrene, and other aromatic hydrocarbons. Here, six ring-hydroxylating dioxygenases (RHDs) were identified in the complete genome ofCycloclasticussp. P1 and were confirmed to be involved in PAH degradation by enzymatic assays. Further, five gene clusters in its genome were identified to be responsible for PAH degradation. Degradation pathways for naphthalene, phenanthrene, and pyrene were elucidated inCycloclasticussp. P1 based on genomic and transcriptomic analysis and characterization of an interconnected metabolic network. The metabolic pathway overlaps in many steps in the degradation of pyrene, phenanthrene, and naphthalene, which were validated by the detection of metabolic intermediates in cultures. This study describes a pyrene degradation pathway forCycloclasticus.Moreover, the study represents the integration of a PAH metabolic network that comprises pyrene, phenanthrene, and naphthalene degradation pathways. Taken together, these results provide a comprehensive investigation of PAH metabolism inCycloclasticus.IMPORTANCEPAHs are ubiquitous in the environment and are carcinogenic compounds and tend to accumulate in food chains due to their low bioavailability and poor biodegradability.Cycloclasticusis an obligate marine PAH degrader and is widespread in marine environments, while the PAH degradation pathways remain unclear. In this report, the degradation pathways for naphthalene, phenanthrene, and pyrene were revealed, and an integrated PAH metabolic network covering pyrene, phenanthrene, and naphthalene was constructed inCycloclasticus. This overlapping network provides streamlined processing of PAHs to intermediates and ultimately to complete mineralization. Furthermore, these results provide an additional context for the prevalence ofCycloclasticusin oil-polluted marine environments and pelagic settings. In conclusion, these analyses provide a useful framework for understanding the cellular processes involved in PAH metabolism in an ecologically important marine bacterium.

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In Salmonella enterica, Ethanolamine Utilization Is Repressed by 1,2-Propanediol To Prevent Detrimental Mixing of Components of Two Different Bacterial Microcompartments

Journal of Bacteriology ◽

10.1128/jb.00215-15 ◽

2015 ◽

Vol 197 (14) ◽

pp. 2412-2421 ◽

Cited By ~ 18

Author(s):

Ryan Sturms ◽

Nicholas A. Streauslin ◽

Shouqiang Cheng ◽

Thomas A. Bobik

Keyword(s):

Protein Interactions ◽

Regulatory Protein ◽

Gene Clusters ◽

Content Type ◽

Bacterial Microcompartments ◽

Shell Protein ◽

Bacterial Microcompartment ◽

Genes Encoding ◽

Shell Proteins ◽

High Conservation

ABSTRACTBacterial microcompartments (MCPs) are a diverse family of protein-based organelles composed of metabolic enzymes encapsulated within a protein shell. The function of bacterial MCPs is to optimize metabolic pathways by confining toxic and/or volatile metabolic intermediates. About 20% of bacteria produce MCPs, and there are at least seven different types. Different MCPs vary in their encapsulated enzymes, but all have outer shells composed of highly conserved proteins containing bacterial microcompartment domains. Many organisms have genes encoding more than one type of MCP, but given the high homology among shell proteins, it is uncertain whether multiple MCPs can be functionally expressed in the same cell at the same time. In these studies, we examine the regulation of the 1,2-propanediol (1,2-PD) utilization (Pdu) and ethanolamine utilization (Eut) MCPs inSalmonella. Studies showed that 1,2-PD (shown to induce the Pdu MCP) represses transcription of the Eut MCP and that the PocR regulatory protein is required. The results indicate that repression of the Eut MCP by 1,2-PD is needed to prevent detrimental mixing of shell proteins from the Eut and Pdu MCPs. Coexpression of both MCPs impaired the function of the Pdu MCP and resulted in the formation of hybrid MCPs composed of Eut and Pdu MCP components. We also show that plasmid-based expression of individual shell proteins from the Eut MCP or the β-carboxysome impaired the function of Pdu MCP. Thus, the high conservation among bacterial microcompartment (BMC) domain shell proteins is problematic for coexpression of the Eut and Pdu MCPs and perhaps other MCPs as well.IMPORTANCEBacterial MCPs are encoded by nearly 20% of bacterial genomes, and almost 40% of those genomes contain multiple MCP gene clusters. In this study, we examine how the regulation of two different MCP systems (Eut and Pdu) is integrated inSalmonella. Our findings indicate that 1,2-PD (shown to induce the Pdu MCP) represses the Eut MCP to prevent detrimental mixing of Eut and Pdu shell proteins. These findings suggest that numerous organisms which produce more than one type of MCP likely need some mechanism to prevent aberrant shell protein interactions.

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Genetic Analysis of Capsular Polysaccharide Synthesis Gene Clusters from All Serotypes of Streptococcus suis: Potential Mechanisms for Generation of Capsular Variation

Applied and Environmental Microbiology ◽

10.1128/aem.03742-12 ◽

2013 ◽

Vol 79 (8) ◽

pp. 2796-2806 ◽

Cited By ~ 58

Author(s):

Masatoshi Okura ◽

Daisuke Takamatsu ◽

Fumito Maruyama ◽

Takashi Nozawa ◽

Ichiro Nakagawa ◽

...

Keyword(s):

Capsular Polysaccharide ◽

Single Gene ◽

Streptococcus Suis ◽

Gene Clusters ◽

Small Scale ◽

Specific Gene ◽

Chromosomal Locus ◽

Content Type ◽

Comprehensive Information ◽

Genes Encoding

ABSTRACTStreptococcus suisstrains are classified into 35 serotypes on the basis of the antigenicity of their capsular polysaccharides (CPs). CP synthesis genes are known to be clustered on the chromosome (cpsgene cluster). The entirecpsgene clusters ofS. suishave so far been sequenced in 15 serotypes and found to be located betweenorfZandaroA. In this study, to provide comprehensive information aboutS. suisCPs, we sequenced the entirecpsgene clusters of the remaining serotypes and analyzed the complete set ofS. suiscpsgene clusters. Among the 35cpsgene clusters, 22 were located betweenorfZandaroA, whereas the other 13 were flanked by other gene(s) on the chromosomes, and the chromosomal locus was classified into five patterns. By clustering analysis, the predicted products ofcpsgenes found in the 35 serotypes were assigned into 291 homology groups, and all serotypes possessed a serotype-specific gene, except for serotypes 1, 2, 1/2, and 14. Because of the presence of genes encoding flippase (wzx) and polymerase (wzy), CPs of all serotypes were thought to be synthesized by the Wzx/Wzy pathway. Our data also implied the possibility of the transfer of the entire or partialcpsgene clusters amongS. suisstrains, as well as the influence of spontaneous mutations in a single gene or a few genes on the antigenicity of some serotypes. Accumulation of these gene transfers and small-scale mutations may have generated the antigenic diversity ofS. suisCPs.

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