scholarly journals Metabolic Pathway Involved in 6-Chloro-2-Benzoxazolinone Degradation by Pigmentiphaga sp. Strain DL-8 and Identification of the Novel Metal-Dependent Hydrolase CbaA

2016 ◽  
Vol 82 (14) ◽  
pp. 4169-4179 ◽  
Author(s):  
Weiliang Dong ◽  
Fei Wang ◽  
Fei Huang ◽  
Yicheng Wang ◽  
Jie Zhou ◽  
...  
Keyword(s):  
Metabolic Pathway ◽  
Specific Activity ◽  
Degradation Mechanism ◽  
Draft Genome ◽  
Proton Donor ◽  
Binding Motif ◽  
Resting Cells ◽  
Content Type ◽  
Peptide Mass ◽  
Donor And Acceptor

ABSTRACT6-Chloro-2-benzoxazolinone (CDHB) is a precursor of herbicide, insecticide, and fungicide synthesis and has a broad spectrum of biological activity.Pigmentiphagasp. strain DL-8 can transform CDHB into 2-amino-5-chlorophenol (2A5CP), which it then utilizes as a carbon source for growth. The CDHB hydrolase (CbaA) was purified from strain DL-8, which can also hydrolyze 2-benzoxazolinone (BOA), 5-chloro-2-BOA, and benzamide. The specific activity of purified CbaA was 5,900 U · mg protein−1for CDHB, withKmandkcatvalues of 0.29 mM and 8,500 s−1, respectively. The optimal pH for purified CbaA was 9.0, the highest activity was observed at 55°C, and the inactive metal-free enzyme could be reactivated by Mg2+, Ni2+, Ca2+, or Zn2+. Based on the results obtained for the CbaA peptide mass fingerprinting and draft genome sequence of strain DL-8,cbaA(encoding 339 amino acids) was cloned and expressed inEscherichia coliBL21(DE3). CbaA shared 18 to 21% identity with some metal-dependent hydrolases of the PF01499 family and contained the signature metal-binding motif Q127XXXQ131XD133XXXH137. The conserved amino acid residues His288 and Glu301 served as the proton donor and acceptor.E. coliBL21(DE3-pET-cbaA) resting cells could transform 0.2 mM CDHB into 2A5CP. The mutant strain DL-8ΔcbaAlost the ability to degrade CDHB but retained the ability to degrade 2A5CP, consistent with strain DL-8. These results indicated thatcbaAwas the key gene responsible for CDHB degradation by strain DL-8.IMPORTANCE2-Benzoxazolinone (BOA) derivatives are widely used as synthetic intermediates and are also an important group of allelochemicals acting in response to tissue damage or pathogen attack in gramineous plants. However, the degradation mechanism of BOA derivatives by microorganisms is not clear. In the present study, we reported the identification of CbaA and metabolic pathway responsible for the degradation of CDHB inPigmentiphagasp. DL-8. This will provide microorganism and gene resources for the bioremediation of the environmental pollution caused by BOA derivatives.

scholarly journals Hydrolase CehA and Monooxygenase CfdC Are Responsible for Carbofuran Degradation inSphingomonassp. Strain CDS-1

2018 ◽  
Vol 84 (16) ◽  
Author(s):  
Xin Yan ◽  
Wen Jin ◽  
Guang Wu ◽  
Wankui Jiang ◽  
Zhangong Yang ◽  
...  
Keyword(s):  
Benzene Ring ◽  
Gene Knockout ◽  
Degradation Mechanism ◽  
Draft Genome ◽  
Open Reading Frames ◽  
Flavin Mononucleotide ◽  
Genetic Determinants ◽  
Content Type ◽  
Degrading Bacteria ◽  
Reading Frames

ABSTRACTCarbofuran, a broad-spectrum systemic insecticide, has been extensively used for approximately 50 years. Diverse carbofuran-degrading bacteria have been described, among which sphingomonads have exhibited an extraordinary ability to catabolize carbofuran; other bacteria can only convert carbofuran to carbofuran phenol, while all carbofuran-degrading sphingomonads can degrade both carbofuran and carbofuran phenol. However, the genetic basis of carbofuran catabolism in sphingomonads has not been well elucidated. In this work, we sequenced the draft genome ofSphingomonassp. strain CDS-1 that can transform both carbofuran and carbofuran phenol but fails to grow on them. On the basis of the hypothesis that the genes involved in carbofuran catabolism are highly conserved among carbofuran-degrading sphingomonads, two such genes,cehACDS-1andcfdCCDS-1, were predicted from the 84 open reading frames (ORFs) that share ≥95% nucleic acid similarities between strain CDS-1 and another sphingomonadNovosphingobiumsp. strain KN65.2 that is able to mineralize the benzene ring of carbofuran. The results of the gene knockout, genetic complementation, heterologous expression, and enzymatic experiments reveal thatcehACDS-1andcfdCCDS-1are responsible for the conversion of carbofuran and carbofuran phenol, respectively, in strain CDS-1. CehACDS-1hydrolyzes carbofuran to carbofuran phenol. CfdCCDS-1, a reduced flavin mononucleotide (FMNH2)- or reduced flavin adenine dinucleotide (FADH2)-dependent monooxygenase, hydroxylates carbofuran phenol at the benzene ring in the presence of NADH, FMN/FAD, and the reductase CfdX. It is worth noting that we found that carbaryl hydrolase CehAAC100, which was previously demonstrated to have no activity toward carbofuran, can actually convert carbofuran to carbofuran phenol, albeit with very low activity.IMPORTANCEDue to the extensive use of carbofuran over the past 50 years, bacteria have evolved catabolic pathways to mineralize this insecticide, which plays an important role in eliminating carbofuran residue in the environment. This study revealed the genetic determinants of carbofuran degradation inSphingomonassp. strain CDS-1. We speculate that the close homologuescehAandcfdCare highly conserved among other carbofuran-degrading sphingomonads and play the same roles as those described here. These findings deepen our understanding of the microbial degradation mechanism of carbofuran and lay a foundation for the better use of microbes to remediate carbofuran contamination.

scholarly journals AmyM, a Novel Maltohexaose-Forming α-Amylase from Corallococcus sp. Strain EGB

2015 ◽  
Vol 81 (6) ◽  
pp. 1977-1987 ◽  
Author(s):  
Zhoukun Li ◽  
Jiale Wu ◽  
Biying Zhang ◽  
Fei Wang ◽  
Xianfeng Ye ◽  
...  
Keyword(s):  
Signal Peptide ◽  
Specific Activity ◽  
Peptide Mass Fingerprinting ◽  
Soluble Starch ◽  
Industrial Applications ◽  
Product Analysis ◽  
Content Type ◽  
E Coli ◽  
Peptide Mass ◽  
Wide Range

ABSTRACTA novel α-amylase, AmyM, was purified from the culture supernatant ofCorallococcussp. strain EGB. AmyM is a maltohexaose-forming exoamylase with an apparent molecular mass of 43 kDa. Based on the results of matrix-assisted laser desorption ionization–time of flight mass spectrometry and peptide mass fingerprinting of AmyM and by comparison to the genome sequence ofCorallococcuscoralloidesDSM 2259, the AmyM gene was identified and cloned intoEscherichia coli.amyMencodes a secretory amylase with a predicted signal peptide of 23 amino acid residues, which showed no significant identity with known and functionally verified amylases.amyMwas expressed inE. coliBL21(DE3) cells with a hexahistidine tag. The signal peptide efficiently induced the secretion of mature AmyM inE. coli. Recombinant AmyM (rAmyM) was purified by Ni-nitrilotriacetic acid (NTA) affinity chromatography, with a specific activity of up to 14,000 U/mg. rAmyM was optimally active at 50°C in Tris-HCl buffer (50 mM; pH 7.0) and stable at temperatures of <50°C. rAmyM was stable over a wide range of pH values (from pH 5.0 to 10.0) and highly tolerant to high concentrations of salts, detergents, and various organic solvents. Its activity toward starch was independent of calcium ions. TheKmandVmaxof recombinant AmyM for soluble starch were 6.61 mg ml−1and 44,301.5 μmol min−1mg−1, respectively. End product analysis showed that maltohexaose accounted for 59.4% of the maltooligosaccharides produced. These characteristics indicate that AmyM has great potential in industrial applications.

scholarly journals Involvement of the Cytochrome P450 System EthBAD in theN-Deethoxymethylation of Acetochlor by Rhodococcus sp. Strain T3-1

2015 ◽  
Vol 81 (6) ◽  
pp. 2182-2188 ◽  
Author(s):  
Fei Wang ◽  
Jie Zhou ◽  
Zhoukun Li ◽  
Weiliang Dong ◽  
Ying Hou ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Resting Cells ◽  
Butyl Ether ◽  
Content Type ◽  
Cytochrome P450 System ◽  
E Coli ◽  
Fast Detection ◽  
Peptide Mass

ABSTRACTAcetochlor [2-chloro-N-(ethoxymethyl)-N-(2-ethyl-6-methylphenyl)-acetamide] is a widely applied herbicide with potential carcinogenic properties.N-Deethoxymethylation is the key step in acetochlor biodegradation.N-Deethoxymethylase is a multicomponent enzyme that catalyzes the conversion of acetochlor to 2′-methyl-6′-ethyl-2-chloroacetanilide (CMEPA). Fast detection of CMEPA by a two-enzyme (N-deethoxymethylase–amide hydrolase) system was established in this research. Based on the fast detection method, a three-component enzyme was purified fromRhodococcussp. strain T3-1 using ammonium sulfate precipitation and hydrophobic interaction chromatography. The molecular masses of the components of the purified enzyme were estimated to be 45, 43, and 11 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Based on the results of peptide mass fingerprint analysis, acetochlorN-deethoxymethylase was identified as a cytochrome P450 system, composed of a cytochrome P450 oxygenase (43-kDa component; EthB), a ferredoxin (45 kDa; EthA), and a reductase (11 kDa; EthD), that is involved in the degradation of methyltert-butyl ether. The gene clusterethABCDwas cloned by PCR amplification and expressed inEscherichia coliBL21(DE3). Resting cells of a recombinantE. colistrain showed deethoxymethylation activity against acetochlor. Subcloning ofethABCDshowed thatethABDexpressed inE. coliBL21(DE3) has the activity of acetochlorN-deethoxymethylase and is capable of converting acetochlor to CMEPA.

scholarly journals Selection of a Bifidobacterium animalis subsp.lactisStrain with a Decreased Ability To Produce Acetic Acid

2012 ◽  
Vol 78 (9) ◽  
pp. 3338-3342 ◽  
Author(s):  
Abelardo Margolles ◽  
Borja Sánchez
Keyword(s):  
Acetic Acid ◽  
Specific Activity ◽  
Skim Milk ◽  
Acetate Kinase ◽  
Resting Cells ◽  
Wild Type ◽  
Kinase Gene ◽  
Content Type ◽  
Bifidobacterium Animalis ◽  
De Man

ABSTRACTWe have characterized a new strain,Bifidobacterium animalissubsp.lactisCECT 7953, obtained by random UV mutagenesis, which produces less acetic acid than the wild type (CECT 7954) in three different experimental settings: De Man-Rogosa-Sharpe broth without sodium acetate, resting cells, and skim milk. Genome sequencing revealed a single Phe-Ser substitution in the acetate kinase gene product that seems to be responsible for the strain's reduced acid production. Accordingly, acetate kinase specific activity was lower in the low acetate producer. Strain CECT 7953 produced less acetate, less ethanol, and more yoghourt-related volatile compounds in skim milk than the wild type did. Thus, CECT 7953 shows promising potential for the development of dairy products fermented exclusively by a bifidobacterial strain.

scholarly journals Draft Genome Sequence of Urease-Producing Pseudorhodobacter sp. Strain E13, Isolated from the Yellow Sea in Gunsan, South Korea

2019 ◽  
Vol 8 (23) ◽  
Author(s):  
Si Chul Kim ◽  
Hyo Jung Lee
Keyword(s):  
South Korea ◽  
Genome Sequence ◽  
Yellow Sea ◽  
Draft Genome ◽  
The Yellow Sea ◽  
Draft Genome Sequence ◽  
Protein Coding ◽  
Coding Sequences ◽  
Gram Negative ◽  
Content Type

Here, we report the draft genome sequence of Pseudorhodobacter sp. strain E13, a Gram-negative, aerobic, nonflagellated, and rod-shaped bacterium which was isolated from the Yellow Sea in South Korea. The assembled genome sequence is 3,878,578 bp long with 3,646 protein-coding sequences in 159 contigs.

scholarly journals Draft Genome Sequences of 12 Mycolicibacterium smegmatis Strains Resistant to Imidazo[1,2-b][1,2,4,5]Tetrazines

2019 ◽  
Vol 8 (16) ◽  
Author(s):  
Aleksey A. Vatlin ◽  
Kirill V. Shur ◽  
Valery N. Danilenko ◽  
Dmitry A. Maslov
Keyword(s):  
Draft Genome ◽  
Transcriptional Repressor ◽  
Antituberculosis Drug ◽  
Genome Sequences ◽  
Content Type ◽  
Drug Candidates

Here, we report 12 draft genome sequences of mutant Mycolicibacterium smegmatis strains resistant to imidazo[1,2-b][1,2,4,5]tetrazines, which are antituberculosis drug candidates. We have identified 7 different mutations in the MSMEG_1380 gene, which encodes the AcrR/TetR_N transcriptional repressor, which may activate efflux-mediated resistance.

scholarly journals Draft Genome Sequences of Three Flavobacterium psychrophilum Strains Isolated from Diseased Ayu (Plecoglossus altivelis altivelis) Caught at Three Sites in the Kagami River in Kochi, Japan

2019 ◽  
Vol 8 (34) ◽  
Author(s):  
Hazuki Yamashita ◽  
Takayuki Wada ◽  
Yusuke Kato ◽  
Takuji Ikeda ◽  
Masayuki Imajoh
Keyword(s):  
Draft Genome ◽  
Psychrophilic Bacterium ◽  
Genome Sequences ◽  
Plecoglossus Altivelis ◽  
Gram Negative ◽  
Content Type ◽  
Flavobacterium Psychrophilum ◽  
Skin Ulcers ◽  
The Family

Flavobacterium psychrophilum is a Gram-negative, psychrophilic bacterium within the family Flavobacteriaceae. Here, we report the draft genome sequences of three F. psychrophilum strains isolated from skin ulcers of diseased ayu caught by tomozuri angling at three sites in the Kagami River in Japan.

scholarly journals Draft Genome Sequences of Escherichia coli Strains FP2 and FP3, Isolated from the Canada Goose (Branta canadensis)

2018 ◽  
Vol 7 (9) ◽  
Author(s):  
Allison L. Denny ◽  
Susan E. Arruda
Keyword(s):  
Escherichia Coli ◽  
Draft Genome ◽  
Branta Canadensis ◽  
Canada Goose ◽  
Genome Sequences ◽  
Content Type

Draft genomes of two strains of Escherichia coli, FP2 and FP3, isolated from the feces of the Canada goose (Branta canadensis), were sequenced. Genome sizes were 5.26 Mb with a predicted G+C content of 50.54% (FP2) and 5.07 Mb with a predicted G+C content of 50.41% (FP3).

scholarly journals Draft Genome Sequence of the Shrimp Pathogen Vibrio parahaemolyticus ST17.P5-S1, Isolated in Peninsular Malaysia

2018 ◽  
Vol 7 (11) ◽  
Author(s):  
Sridevi Devadas ◽  
Subha Bhassu ◽  
Tze Chiew Christie Soo ◽  
Fatimah M. Yusoff ◽  
Mohamed Shariff
Keyword(s):  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Vibrio Parahaemolyticus ◽  
East Coast ◽  
Draft Genome ◽  
Antibiotic Resistance Genes ◽  
Peninsular Malaysia ◽  
Penaeus Vannamei ◽  
Content Type ◽  
Acute Hepatopancreatic Necrosis Disease

We sequenced the genome of Vibrio parahaemolyticus strain ST17.P5-S1, isolated from Penaeus vannamei cultured in the east coast of Peninsular Malaysia. The strain contains several antibiotic resistance genes and a plasmid encoding the Photorhabdus insect-related (Pir) toxin-like genes, pirAvp and pirBvp, associated with acute hepatopancreatic necrosis disease (AHPND).

scholarly journals Draft Genome Sequence of a Vibrio harveyi Strain Associated with Vibriosis in Pacific White Shrimp (Litopenaeus vannamei)

2017 ◽  
Vol 5 (7) ◽  
Author(s):  
Emille Moreno ◽  
Marci Parks ◽  
Lee J. Pinnell ◽  
James J. Tallman ◽  
Jeffrey W. Turner
Keyword(s):  
Genome Sequence ◽  
Litopenaeus Vannamei ◽  
Vibrio Harveyi ◽  
Draft Genome ◽  
Penaeid Shrimp ◽  
Pacific White Shrimp ◽  
Draft Genome Sequence ◽  
White Shrimp ◽  
Content Type ◽  
Future Studies

ABSTRACT Vibrio harveyi is a Gram-negative bacterium associated with vibriosis in penaeid shrimp. Here, we report the draft genome sequence of a V. harveyi strain isolated from Pacific white shrimp (Litopenaeus vannamei) during a vibriosis outbreak. The availability of this genome will aid future studies of vibriosis in shrimp aquaculture.

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