scholarly journals DdvK, a Novel Major Facilitator Superfamily Transporter Essential for 5,5′-Dehydrodivanillate Uptake bySphingobiumsp. Strain SYK-6

2018 ◽  
Vol 84 (20) ◽  
Author(s):  
Kosuke Mori ◽  
Koh Niinuma ◽  
Masaya Fujita ◽  
Naofumi Kamimura ◽  
Eiji Masai
Keyword(s):  
Value Added ◽  
Major Facilitator Superfamily ◽  
Amino Acid Sequence Similarity ◽  
Aromatic Acids ◽  
Transporter Gene ◽  
Content Type ◽  
Biomass Resource ◽  
Major Facilitator ◽  
Mfs Transporter

ABSTRACTThe microbial conversion of lignin-derived aromatics is a promising strategy for the industrial utilization of this large biomass resource. However, efficient application requires an elucidation of the relevant transport and catabolic pathways. InSphingobiumsp. strain SYK-6, most of the enzyme genes involved in 5,5′-dehydrodivanillate (DDVA) catabolism have been characterized, but the transporter has not yet been identified. Here, we identified SLG_07710 (ddvK) and SLG_07780 (ddvR), genes encoding a putative major facilitator superfamily (MFS) transporter and MarR-type transcriptional regulator, respectively. AddvKmutant of SYK-6 completely lost the capacity to grow on and convert DDVA. DdvR repressed the expression of the DDVAO-demethylase oxygenase component gene (ligXa), while DDVA acted as the gene inducer. A DDVA uptake assay was developed by employing this DdvR-controlledligXatranscriptional regulatory system. ASphingobium japonicumUT26S transformant expressingddvKacquired DDVA uptake capacity, indicating thatddvKencodes the DDVA transporter. DdvK, probably requiring the proton motive force, was suggested to be a novel MFS transporter on the basis of the amino acid sequence similarity. Subsequently, we evaluated the effects ofddvKoverexpression on the production of the DDVA metabolite 2-pyrone-4,6-dicarboxylate (PDC), a building block of functional polymers. A SYK-6 mutant of the PDC hydrolase gene (ligI) cultured in DDVA accumulated PDC via 5-carboxyvanillate and grew by utilizing 4-carboxy-2-hydroxypenta-2,4-dienoate. The introduction of addvK-expression plasmid into aligImutant increased the growth rate in DDVA and the amounts of DDVA converted and PDC produced after 48 h by 1.35- and 1.34-fold, respectively. These results indicate that enhanced transporter gene expression can improve metabolite production from lignin derivatives.IMPORTANCEThe bioengineering of bacteria to selectively transport and metabolize natural substrates into specific metabolites is a valuable strategy for industrial-scale chemical production. The uptake of many substrates into cells requires specific transport systems, and so the identification and characterization of transporter genes are essential for industrial applications. A number of bacterial major facilitator superfamily transporters of aromatic acids have been identified and characterized, but many transporters of lignin-derived aromatic acids remain unidentified. The efficient conversion of lignin, an abundant but unutilized aromatic biomass resource, to value-added metabolites using microbial catabolism requires the characterization of transporters for lignin-derived aromatics. In this study, we identified the transporter gene responsible for the uptake of 5,5′-dehydrodivanillate, a lignin-derived biphenyl compound, inSphingobiumsp. strain SYK-6. In addition to characterizing its function, we applied this transporter gene to the production of a value-added metabolite from 5,5′-dehydrodivanillate.

scholarly journals Enhanced Efflux Pump Expression in Candida Mutants Results in Decreased Manogepix Susceptibility

2020 ◽  
Vol 64 (5) ◽  
Author(s):  
Sean D. Liston ◽  
Luke Whitesell ◽  
Mili Kapoor ◽  
Karen Joy Shaw ◽  
Leah E. Cowen
Keyword(s):  
Fungal Pathogens ◽  
Candida Parapsilosis ◽  
Efflux Pump ◽  
Major Facilitator Superfamily ◽  
Transporter Gene ◽  
Transcription Factor Gene ◽  
Content Type ◽  
Atp Binding Cassette Transporter ◽  
Major Facilitator Superfamily Transporter ◽  
Major Facilitator

ABSTRACT Manogepix is a broad-spectrum antifungal agent that inhibits glycosylphosphatidylinositol (GPI) anchor biosynthesis. Using whole-genome sequencing, we characterized two efflux-mediated mechanisms in the fungal pathogens Candida albicans and Candida parapsilosis that resulted in decreased manogepix susceptibility. In C. albicans, a gain-of-function mutation in the transcription factor gene ZCF29 activated expression of ATP-binding cassette transporter genes CDR11 and SNQ2. In C. parapsilosis, a mitochondrial deletion activated expression of the major facilitator superfamily transporter gene MDR1.

scholarly journals ThemelREDCAOperon Encodes a Utilization System for the Raffinose Family of Oligosaccharides inBacillus subtilis

2019 ◽  
Vol 201 (15) ◽  
Author(s):  
Kambiz Morabbi Heravi ◽  
Hildegard Watzlawick ◽  
Josef Altenbuchner
Keyword(s):  
Bacillus Subtilis ◽  
Transcriptional Start Site ◽  
Major Facilitator Superfamily ◽  
Atp Binding ◽  
Carbohydrate Utilization ◽  
Content Type ◽  
Storage Organs ◽  
Major Facilitator ◽  
Atp Binding Cassette

ABSTRACTBacillus subtilisis a heterotrophic soil bacterium that hydrolyzes different polysaccharides mainly found in the decomposed plants. These carbohydrates are mainly cellulose, hemicellulose, and the raffinose family of oligosaccharides (RFOs). RFOs are soluble α-galactosides, such as raffinose, stachyose, and verbascose, that rank second only after sucrose in abundance. Genome sequencing and transcriptome analysis ofB. subtilisindicated the presence of a putative α-galactosidase-encoding gene (melA) located in themsmRE-amyDC-melAoperon. Characterization of the MelA protein showed that it is a strictly Mn2+- and NAD+-dependent α-galactosidase able to hydrolyze melibiose, raffinose, and stachyose. Transcription of themsmER-amyDC-melAoperon is under control of a σA-type promoter located upstream ofmsmR(PmsmR), which is negatively regulated by MsmR. The activity of PmsmRwas induced in the presence of melibiose and raffinose. MsmR is a transcriptional repressor that binds to two binding sites at PmsmRlocated upstream of the −35 box and downstream of the transcriptional start site. MsmEX-AmyCD forms an ATP-binding cassette (ABC) transporter that probably transports melibiose into the cell. SincemsmRE-amyDC-melAis a melibiose utilization system, we renamed the operonmelREDCA.IMPORTANCEBacillus subtilisutilizes different polysaccharides produced by plants. These carbohydrates are primarily degraded by extracellular hydrolases, and the resulting oligo-, di-, and monosaccharides are transported into the cytosol via phosphoenolpyruvate-dependent phosphotransferase systems (PTS), major facilitator superfamily, and ATP-binding cassette (ABC) transporters. In this study, a new carbohydrate utilization system ofB. subtilisresponsible for the utilization of α-galactosides of the raffinose family of oligosaccharides (RFOs) was investigated. RFOs are synthesized from sucrose in plants and are mainly found in the storage organs of plant leaves. Our results revealed the modus operandi of a new carbohydrate utilization system inB. subtilis.

scholarly journals Genome Sequence of a Staphylococcus xylosus Clinical Isolate, Strain SMA0341-04 (UGA5), from Siaya County Referral Hospital in Siaya, Kenya

2019 ◽  
Vol 8 (16) ◽  
Author(s):  
Qiuying Cheng ◽  
Gary Xie ◽  
Hajnalka Daligault ◽  
Karen Davenport ◽  
Cheryl Gleasner ◽  
...  
Keyword(s):  
Clinical Isolate ◽  
Genome Sequence ◽  
Major Facilitator Superfamily ◽  
Referral Hospital ◽  
Content Type ◽  
Staphylococcus Xylosus ◽  
Major Facilitator ◽  
Mfs Transporter

We report here the genome sequence of a Staphylococcus xylosus clinical isolate, strain SMA0341-04 (UGA5), which contains one chromosome and at least one plasmid. Notably, strain SMA0341-04 (UGA5) contains the tetracycline efflux major facilitator superfamily (MFS) transporter (tetK) gene.

scholarly journals Missense Mutations in CytochromecMaturation Genes Provide New Insights into Rhodobacter capsulatus cbb3-Type CytochromecOxidase Biogenesis

2012 ◽  
Vol 195 (2) ◽  
pp. 261-269 ◽  
Author(s):  
Seda Ekici ◽  
Xinpei Jiang ◽  
Hans-Georg Koch ◽  
Fevzi Daldal
Keyword(s):  
Rhodobacter Capsulatus ◽  
Major Facilitator Superfamily ◽  
Missense Mutations ◽  
Content Type ◽  
Knockout Mutants ◽  
Major Facilitator ◽  
Copper Oxidase ◽  
First Time ◽  
Derivatives Of

ABSTRACTTheRhodobacter capsulatus cbb3-type cytochromecoxidase (cbb3-Cox) belongs to the heme-copper oxidase superfamily, and its subunits are encoded by theccoNOQPoperon. Biosynthesis of this enzyme is complex and needs dedicated biogenesis genes (ccoGHIS). It also relies on thec-type cytochrome maturation (Ccm) process, which requires theccmABCDEFGHIgenes, because two of thecbb3-Cox subunits (CcoO and CcoP) arec-type cytochromes. Recently, we reported that mutants lacking CcoA, a major facilitator superfamily type transporter, produce very small amounts ofcbb3-Cox unless the growth medium is supplemented with copper. In this work, we isolated “Cu-unresponsive” derivatives of accoAdeletion strain that exhibited nocbb3-Cox activity even upon Cu supplementation. Molecular characterization of these mutants revealed missense mutations in theccmAorccmFgene, required for the Ccm process. As expected, Cu-unresponsive mutants lacked the CcoO and CcoP subunits due to Ccm defects, but remarkably, they contained the CcoN subunit ofcbb3-Cox. Subsequent construction and examination of singleccmknockout mutants demonstrated that membrane insertion and stability of CcoN occurred in the absence of the Ccm process. Moreover, while theccmknockout mutants were completely incompetent for photosynthesis, the Cu-unresponsive mutants grew photosynthetically at lower rates and produced smaller amounts of cytochromesc1andc2than did a wild-type strain due to their restricted Ccm capabilities. These findings demonstrate that different levels of Ccm efficiency are required for the production of variousc-type cytochromes and reveal for the first time that maturation of the heme-Cu-containing subunit CcoN ofR. capsulatus cbb3-Cox proceeds independently of that of thec-type cytochromes during the biogenesis of this enzyme.

scholarly journals Draft Genome Sequences of Two Staphylococcus warneri Clinical Isolates, Strains SMA0023-04 (UGA3) and SMA0670-05 (UGA28), from Siaya County Referral Hospital, Siaya, Kenya

2019 ◽  
Vol 8 (15) ◽  
Author(s):  
Gary Xie ◽  
Qiuying Cheng ◽  
Hajnalka Daligault ◽  
Karen Davenport ◽  
Cheryl Gleasner ◽  
...  
Keyword(s):  
Draft Genome ◽  
Macrolide Resistance ◽  
Major Facilitator Superfamily ◽  
Clinical Isolates ◽  
Beta Lactamase ◽  
Genome Sequences ◽  
Content Type ◽  
Staphylococcus Warneri ◽  
Major Facilitator ◽  
Mfs Transporter

We report the complete draft genome sequences of two Staphylococcus warneri clinical isolates, strains SMA0023-04 (UGA3) and SMA0670-05 (UGA28), each of which contains one chromosome and at least one plasmid. Isolate SMA0023-04 (UGA3) contains tetracycline efflux major facilitator superfamily (MFS) transporter (tetK), macrolide resistance (msrC and mphC), and beta-lactamase (blaZ) genes on its plasmids.

scholarly journals Functional Characterization of AbaQ, a Novel Efflux Pump Mediating Quinolone Resistance inAcinetobacter baumannii

2018 ◽  
Vol 62 (9) ◽  
Author(s):  
María Pérez-Varela ◽  
Jordi Corral ◽  
Jesús Aranda ◽  
Jordi Barbé
Keyword(s):  
Acinetobacter Baumannii ◽  
Antimicrobial Agents ◽  
Efflux Pump ◽  
Functional Characterization ◽  
Multidrug Resistant ◽  
Quinolone Resistance ◽  
Nosocomial Pathogen ◽  
Content Type ◽  
Mfs Transporter

ABSTRACTAcinetobacter baumanniihas emerged as an important multidrug-resistant nosocomial pathogen. In previous work, we identified a putative MFS transporter, AU097_RS17040, involved in the pathogenicity ofA. baumannii(M. Pérez-Varela, J. Corral, J. A. Vallejo, S. Rumbo-Feal, G. Bou, J. Aranda, and J. Barbé, Infect Immun 85:e00327-17, 2017,https://doi.org/10.1128/IAI.00327-17). In this study, we analyzed the susceptibility to diverse antimicrobial agents ofA. baumanniicells defective in this transporter, referred to as AbaQ. Our results showed that AbaQ is mainly involved in the extrusion of quinolone-type drugs inA. baumannii.

scholarly journals Bacterial Catabolism of β-Hydroxypropiovanillone and β-Hydroxypropiosyringone Produced in the Reductive Cleavage of Arylglycerol-β-Aryl Ether in Lignin

2018 ◽  
Vol 84 (7) ◽  
Author(s):  
Yudai Higuchi ◽  
Shogo Aoki ◽  
Hiroki Takenami ◽  
Naofumi Kamimura ◽  
Kenji Takahashi ◽  
...  
Keyword(s):  
Value Added ◽  
Lignin Biodegradation ◽  
Gene Products ◽  
Aryl Ether ◽  
Ether Linkage ◽  
Content Type ◽  
Catabolic Genes ◽  
Genes Encoding ◽  
Stereospecific Reaction

ABSTRACTSphingobiumsp. strain SYK-6 converts four stereoisomers of arylglycerol-β-guaiacyl ether into achiral β-hydroxypropiovanillone (HPV) via three stereospecific reaction steps. Here, we determined the HPV catabolic pathway and characterized the HPV catabolic genes involved in the first two steps of the pathway. In SYK-6 cells, HPV was oxidized to vanilloyl acetic acid (VAA) via vanilloyl acetaldehyde (VAL). The resulting VAA was further converted into vanillate through the activation of VAA by coenzyme A. A syringyl-type HPV analog, β-hydroxypropiosyringone (HPS), was also catabolized via the same pathway. SLG_12830 (hpvZ), which belongs to the glucose-methanol-choline oxidoreductase family, was isolated as the HPV-converting enzyme gene. AnhpvZmutant completely lost the ability to convert HPV and HPS, indicating thathpvZis essential for the conversion of both the substrates. HpvZ produced inEscherichia colioxidized both HPV and HPS and other 3-phenyl-1-propanol derivatives. HpvZ localized to both the cytoplasm and membrane of SYK-6 and used ubiquinone derivatives as electron acceptors. Thirteen gene products of the 23 aldehyde dehydrogenase (ALDH) genes in SYK-6 were able to oxidize VAL into VAA. Mutant analyses suggested that multiple ALDH genes, including SLG_20400, contribute to the conversion of VAL. We examined whether the genes encoding feruloyl-CoA synthetase (ferA) and feruloyl-CoA hydratase/lyase (ferBandferB2) are involved in the conversion of VAA. Only FerA exhibited activity toward VAA; however, disruption offerAdid not affect VAA conversion. These results indicate that another enzyme system is involved in VAA conversion.IMPORTANCECleavage of the β-aryl ether linkage is the most essential process in lignin biodegradation. Although the bacterial β-aryl ether cleavage pathway and catabolic genes have been well documented, there have been no reports regarding the catabolism of HPV or HPS, the products of cleavage of β-aryl ether compounds. HPV and HPS have also been found to be obtained from lignin by chemoselective catalytic oxidation by 2,3-dichloro-5,6-dicyano-1,4-benzoquinone/tert-butyl nitrite/O2, followed by cleavage of the β-aryl ether with zinc. Therefore, value-added chemicals are expected to be produced from these compounds. In this study, we determined the SYK-6 catabolic pathways for HPV and HPS and identified the catabolic genes involved in the first two steps of the pathways. Since SYK-6 catabolizes HPV through 2-pyrone-4,6-dicarboxylate, which is a building block for functional polymers, characterization of HPV catabolism is important not only for understanding the bacterial lignin catabolic system but also for lignin utilization.

scholarly journals Characterization of Glucose-Specific Catabolite Repression-Resistant Mutants of Bacillus subtilis: Identification of a Novel Hexose:H+ Symporter

1998 ◽  
Vol 180 (3) ◽  
pp. 498-504 ◽  
Author(s):  
Ian T. Paulsen ◽  
Sylvie Chauvaux ◽  
Peter Choi ◽  
Milton H. Saier
Keyword(s):  
Bacillus Subtilis ◽  
Catabolite Repression ◽  
Genetic Background ◽  
Insertional Mutagenesis ◽  
Sequence Similarity ◽  
Major Facilitator Superfamily ◽  
Phosphotransferase System ◽  
E Coli ◽  
Major Facilitator

ABSTRACT Insertional mutagenesis was conducted on Bacillus subtilis cells to screen for mutants resistant to catabolite repression. Three classes of mutants that were resistant to glucose-promoted but not mannitol-promoted catabolite repression were identified. Cloning and sequencing of the mutated genes revealed that the mutations occurred in the structural genes for (i) enzyme II of the phosphoenolpyruvate-glucose phosphotransferase (PtsG), (ii) antiterminator GlcT, which controls PtsG synthesis, and (iii) a previously uncharacterized carrier of the major facilitator superfamily, which we have designated GlcP. The last protein exhibits greatest sequence similarity to the fucose:H+ symporter ofEscherichia coli and the glucose/galactose:H+symporter of Brucella abortus. In a wild-type B. subtilis genetic background, theglcP::Tn10 mutation (i) partially but specifically relieved glucose- and sucrose-promoted catabolite repression, (ii) reduced the growth rate in minimal glucose medium, and (iii) reduced rates of [14C]glucose and [14C]methyl α-glucoside uptake. In a Δptsgenetic background no phenotype was observed, suggesting that expression of the glcP gene required a functional phosphotransferase system. When overproduced in a Δptsmutant of E. coli, GlcP could be shown to specifically transport glucose, mannose, 2-deoxyglucose and methyl α-glucoside with low micromolar affinities. Accumulation of the nonmetabolizable glucose analogs was demonstrated, and inhibitor studies suggested a dependency on the proton motive force. We conclude that B. subtilis possesses at least two distinct routes of glucose entry, both of which contribute to the phenomenon of catabolite repression.

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