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Dehalogenase

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https://www.readbyqxmd.com/read/28282255/effect-of-zn-2-on-halohydrin-dehalogenase-expression-and-accumulation-through-multi-parameter-correlation-research-with-escherichia-coli-p84a-mc1061
#1
Xiao Li, Yu-Di Luo, Dong-Rui Pan, Xiao-Dan Shi, Ya-Li Tan, Zhi-Hong Li
Using 5 Zn(2+) supplementation strategies in a 50 L batch bioreactor named FUS-50L(A), possible correlations among Zn(2+) content and addition timing, physiologic activity (PA), halohydrin dehalogenase (HheC) accumulation of Escherichia coli P84A/MC1061 were systematically investigated. First, Zn(2+) was confirmed as the significant factor, and its optimal concentration for HheC expression was 3.87 mg/L through fermentation experiments in shaking flasks. Second, based on experimental results from the different strategies, it was found that PA, nutrient consumption rate (NCR) and specific growth rate (μ) for E...
March 10, 2017: Bioengineered
https://www.readbyqxmd.com/read/28265269/marine-derived-biocatalysts-importance-accessing-and-application-in-aromatic-pollutant-bioremediation
#2
REVIEW
Efstratios Nikolaivits, Maria Dimarogona, Nikolas Fokialakis, Evangelos Topakas
The aim of the present review is to highlight the potential use of marine biocatalysts (whole cells or enzymes) as an alternative bioprocess for the degradation of aromatic pollutants. Firstly, information about the characteristics of the still underexplored marine environment and the available scientific tools used to access novel marine-derived biocatalysts is provided. Marine-derived enzymes, such as dioxygenases and dehalogenases, and the involved catalytic mechanisms for the degradation of aromatic and halogenated compounds, are presented, with the purpose of underpinning their potential use in bioremediation...
2017: Frontiers in Microbiology
https://www.readbyqxmd.com/read/28251763/a-13-week-research-based-biochemistry-laboratory-curriculum
#3
Scott T Lefurgy, Emily C Mundorff
Here, we present a 13-week research-based biochemistry laboratory curriculum designed to provide the students with the experience of engaging in original research while introducing foundational biochemistry laboratory techniques. The laboratory experience has been developed around the directed evolution of an enzyme chosen by the instructor, with mutations designed by the students. Ideal enzymes for this curriculum are able to be structurally modeled, solubly expressed, and monitored for activity by UV/Vis spectroscopy, and an example curriculum for haloalkane dehalogenase is given...
March 2, 2017: Biochemistry and Molecular Biology Education
https://www.readbyqxmd.com/read/28238079/development-and-application-of-a-rapid-user-friendly-and-inexpensive-method-to-detect-dehalococcoides-sp-reductive-dehalogenase-genes-from-groundwater
#4
Yogendra H Kanitkar, Robert D Stedtfeld, Paul B Hatzinger, Syed A Hashsham, Alison M Cupples
TaqMan probe-based quantitative polymerase chain reaction (qPCR) specific to the biomarker reductive dehalogenase (RDase) genes is a widely accepted molecular biological tool (MBT) for determining the abundance of Dehalococcoides sp. in groundwater samples from chlorinated solvent-contaminated sites. However, there are significant costs associated with this MBT. In this study, we describe an approach that requires only low-cost laboratory equipment (a bench top centrifuge and a water bath) and requires less time and resources compared to qPCR...
February 25, 2017: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28191945/engineered-surface-immobilized-enzyme-that-retains-high-levels-of-catalytic-activity-in-air
#5
Somayesadat Badieyan, Qiuming Wang, Xingquan Zou, Yaoxin Li, Maggie Herron, Nicholas L Abbott, Zhan Chen, E Neil G Marsh
In the absence of aqueous buffer, most enzymes retain little or no activity; however, "water-free" enzymes would have many diverse applications. Here, we describe the chemically precise immobilization of an enzyme on an engineered surface designed to support catalytic activity in air at ambient humidity. Covalent immobilization of haloalkane dehalogenase on a surface support displaying poly(sorbitol methacrylate) chains resulted in ∼40-fold increase in activity over lyophilized enzyme powders for the gas-phase dehalogenation of 1-bromopropane...
February 16, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28191552/a-common-mechanism-for-coenzyme-cobalamin-dependent-reductive-dehalogenases
#6
Linus O Johannissen, David Leys, Sam Hay
Distinct mechanisms have been proposed for the biological dehalogenation catalyzed by cobalamin-dependent enzymes, with two recent crystallographic studies suggesting different mechanisms based on the observed interaction between the organohalide substrate and cobalamin. In one case, involving an aromatic dibromide substrate in NpRdhA, a novel Co(II)-Br interaction was observed using EPR, suggesting a mechanism involving a [CoXR] adduct. However, in the case of trichloroethylene in PceA, a significantly longer Co-Cl distance was observed in X-ray crystal structures, suggesting a dissociative electron transfer mechanism...
February 13, 2017: Physical Chemistry Chemical Physics: PCCP
https://www.readbyqxmd.com/read/28165015/enhancing-the-biocatalytic-manufacture-of-the-key-intermediate-of-atorvastatin-by-focused-directed-evolution-of-halohydrin-dehalogenase
#7
Yu Luo, Yangzi Chen, Hongmin Ma, ZhenHua Tian, Yeqi Zhang, Jian Zhang
Halohydrin dehalogenases (HHDHs) are biocatalytically interesting enzymes due to their ability to form C-C, C-N, C-O, and C-S bonds. One of most important application of HHDH was the protein engineering of HheC (halohydrin dehalogenase from Agrobacterium radiobacter AD1) for the industrial manufacturing of ethyl (R)-4-cyano-3-hydroxybutanoate (HN), a key chiral synthon of a cholesterol-lowering drug of atorvastatin. During our development of an alternative, more efficient and economic route for chemo-enzymatic preparation of the intermediate of atorvastatin, we found that the HheC2360 previously reported for HN manufacture, had insufficient activity for the cyanolysis production of tert-butyl (3 R,5 S)-6-cyano-3,5-dihydroxyhexanoate (A7)...
February 6, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28104837/the-role-of-dimer-asymmetry-and-protomer-dynamics-in-enzyme-catalysis
#8
Tae Hun Kim, Pedram Mehrabi, Zhong Ren, Adnan Sljoka, Christopher Ing, Alexandr Bezginov, Libin Ye, Régis Pomès, R Scott Prosser, Emil F Pai
Freeze-trapping x-ray crystallography, nuclear magnetic resonance, and computational techniques reveal the distribution of states and their interconversion rates along the reaction pathway of a bacterial homodimeric enzyme, fluoroacetate dehalogenase (FAcD). The crystal structure of apo-FAcD exhibits asymmetry around the dimer interface and cap domain, priming one protomer for substrate binding. This asymmetry is dynamically averaged through conformational exchange on a millisecond time scale. During catalysis, the protomer conformational exchange rate becomes enhanced, the empty protomer exhibits increased local disorder, and water egresses...
January 20, 2017: Science
https://www.readbyqxmd.com/read/28075122/acetylene-fuels-tce-reductive-dechlorination-by-defined-dehalococcoides-pelobacter-consortia
#9
Xinwei Mao, Ronald S Oremland, Tong Liu, Sara Gushgari, Abigail A Landers, Shaun M Baesman, Lisa Alvarez-Cohen
Acetylene (C2H2) can be generated in contaminated groundwater sites as a consequence of chemical degradation of trichloroethene (TCE) by in situ minerals, and C2H2 is known to inhibit bacterial dechlorination. In this study, we show that while high C2H2 (1.3 mM) concentrations reversibly inhibit reductive dechlorination of TCE by Dehalococcoides mccartyi isolates as well as enrichment cultures containing D. mccartyi sp., low C2H2 (0.4 mM) concentrations do not inhibit growth or metabolism of D. mccartyi. Cocultures of Pelobacter SFB93, a C2H2-fermenting bacterium, with D...
February 3, 2017: Environmental Science & Technology
https://www.readbyqxmd.com/read/28074221/exploring-the-thermostable-properties-of-halohydrin-dehalogenase-from-agrobacterium-radiobacter-ad1-by-a-combinatorial-directed-evolution-strategy
#10
Zhiyun Wu, Wenfeng Deng, Yapei Tong, Qian Liao, Dongmin Xin, Huashun Yu, Juan Feng, Lixia Tang
As a crucial factor for biocatalysts, protein thermostability often arises from a combination of factors that are often difficult to rationalize. In this work, the thermostable nature of halohydrin dehalogenase from Agrobacterium radiobacter AD1 (HheC) was systematically explored using a combinatorial directed evolution approach. For this, a mutagenesis library of HheC mutants was first constructed using error-prone PCR with low mutagenesis frequency. After screening approximately 2000 colonies, six mutants with eight mutation sites were obtained...
January 10, 2017: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28063652/molecular-and-carbon-isotopic-characterization-of-an-anaerobic-stable-enrichment-culture-containing-dehalobacterium-sp-during-dichloromethane-fermentation
#11
Alba Trueba-Santiso, Eloi Parladé, Mònica Rosell, Marc Lliros, Siti Hatijah Mortan, Maira Martínez-Alonso, Nuria Gaju, Lucía Martín-González, Teresa Vicent, Ernest Marco-Urrea
Biodegradation of dichloromethane (DCM) under reducing conditions is of major concern due to its widespread detection in contaminated groundwaters. Here, we report an anaerobic enrichment culture derived from a membrane bioreactor operating in an industrial wastewater treatment plant, capable of fermenting DCM and the brominated analogue dibromomethane (DBM). Comparative analysis of bacterial 16S rDNA-DGGE profiles from fresh liquid medium inoculated with single colonies picked from serial dilution-to-extinction agar vials showed that cultures degrading DCM contained a predominant band belonging to Dehalobacterium, however this band was absent in cultures unable to degrade DCM...
January 4, 2017: Science of the Total Environment
https://www.readbyqxmd.com/read/28002948/survival-of-vinyl-chloride-respiring-dehalococcoides-mccartyi-under-long-term-electron-donor-limitation
#12
Koshlan Mayer-Blackwell, Mohammad F Azizian, Jennifer K Green, Alfred M Spormann, Lewis Semprini
In anoxic groundwater aquifers, the long-term survival of Dehalococcoides mccartyi populations expressing the gene vcrA (or bvcA) encoding reductive vinyl chloride dehalogenases are important to achieve complete dechlorination of tetrachloroethene (PCE) and trichloroethene (TCE) to nonchlorinated ethene. The absence or inactivity of vcrA-containing Dehalococcoides results in the accumulation of the harmful chlorinated intermediates dichloroethene (DCE) and vinyl chloride (VC). Although vcrA-containing Dehalococcoides subpopulations depend on synergistic interaction with other organohalide-respiring populations generating their metabolic electron acceptors (DCE and VC), their survival requires successful competition for electron donor within the entire organohalide-respiring microbial community...
January 17, 2017: Environmental Science & Technology
https://www.readbyqxmd.com/read/27999093/a-robust-cosolvent-compatible-halohydrin-dehalogenase-by-computational-library-design
#13
Hesam Arabnejad, Marco Dal Lago, Peter A Jekel, Robert J Floor, Andy-Mark W H Thunnissen, Anke C Terwisscha van Scheltinga, Hein J Wijma, Dick B Janssen
To improve the applicability of halohydrin dehalogenase as a catalyst for reactions in the presence of organic cosolvents, we explored a computational library design strategy (Framework for Rapid Enzyme Stabilization by Computational libraries) that involves discovery and in silico evaluation of stabilizing mutations. Energy calculations, disulfide bond predictions and molecular dynamics simulations identified 218 point mutations and 35 disulfide bonds with predicted stabilizing effects. Experiments confirmed 29 stabilizing point mutations, most of which were located in two distinct regions, whereas introduction of disulfide bonds was not effective...
December 19, 2016: Protein Engineering, Design & Selection: PEDS
https://www.readbyqxmd.com/read/27907199/identification-heterologous-expression-and-functional-characterization-of-bacillus-subtilis-yutf-a-had-superfamily-5-nucleotidase-with-broad-substrate-specificity
#14
Natalia P Zakataeva, Dmitriy V Romanenkov, Yuliya R Yusupova, Victoria S Skripnikova, Takayuki Asahara, Sergey V Gronskiy
5'-nucleotidases (EC 3.1.3.5) catalyze the hydrolytic dephosphorylation of 5'-ribonucleotides and 5'-deoxyribonucleotides as well as complex nucleotides, such as uridine 5'-diphosphoglucose (UDP-glucose), nicotinamide adenine dinucleotide and flavin adenine dinucleotide, to their corresponding nucleosides plus phosphate. These enzymes have been found in diverse species in intracellular and membrane-bound, surface-localized forms. Soluble forms of 5'-nucleotidases belong to the ubiquitous haloacid dehalogenase superfamily (HADSF) and have been shown to be involved in the regulation of nucleotide, nucleoside and nicotinamide adenine dinucleotide (NAD+) pools...
2016: PloS One
https://www.readbyqxmd.com/read/27863020/kinetics-of-binding-of-fluorescent-ligands-to-enzymes-with-engineered-access-tunnels
#15
Shubhangi Kaushik, Zbynek Prokop, Jiri Damborsky, Radka Chaloupkova
Molecular recognition mechanisms and kinetics of binding of ligands to buried active sites via access tunnels are not well understood. Fluorescence polarization enables rapid and non-destructive real-time quantification of the association between small fluorescent ligands and large biomolecules. In this study, we describe analysis of binding kinetics of fluorescent ligands resembling linear halogenated alkanes to haloalkane dehalogenases. Dehalogenases possess buried active sites connected to the surrounding solvent by access tunnels...
November 9, 2016: FEBS Journal
https://www.readbyqxmd.com/read/27856620/genomic-characterization-of-dehalococcoides-mccartyi-strain-11a5-reveals-a-circular-extrachromosomal-genetic-element-and-a-new-tetrachloroethene-reductive-dehalogenase-gene
#16
Siyan Zhao, Chang Ding, Jianzhong He
Dehalococcoides mccartyi exhibits versatile capabilities to respire halogenated compounds under anaerobic conditions. In this study, we report the assembly and annotation of the complete genome of a chloroethene dechlorinating D. mccartyi strain 11a5. Bearing a 1,461,973 base-pair chromosome, strain 11a5 distinguishes itself from other D. mccartyi strains by possessing a 5,940 base-pair circular extrachromosomal genetic element which contains a reductive dehalogenase homolog. The whole genome of strain 11a5 harbors 31 putative reductive dehalogenase genes...
November 16, 2016: FEMS Microbiology Ecology
https://www.readbyqxmd.com/read/27836849/identification-of-a-new-phosphatase-enzyme-potentially-involved-in-the-sugar-phosphate-stress-response-in-pseudomonas-fluorescens
#17
Susan Maleki, Radka Hrudikova, Sergey B Zotchev, Helga Ertesvåg
The alginate-producing bacterium Pseudomonas fluorescens utilizes the Entner-Doudoroff (ED) and pentose phosphate (PP) pathways to metabolize fructose, since the upper part of its Embden-Meyerhof-Parnas pathway is defective. Our previous study indicated that perturbation of the central carbon metabolism by diminishing glucose-6-phosphate dehydrogenase activity could lead to sugar phosphate stress when P. fluorescens was cultivated on fructose. In the present study, we demonstrate that PFLU2693, annotated as a haloacid dehalogenase-like enzyme, is a new sugar phosphate phosphatase, now designated Spp, which is able to dephosphorylate a range of phosphate substrates, including glucose 6-phosphate and fructose 6-phosphate, in vitro The effect of spp overexpression on growth and alginate production was investigated using both the wild type and several mutant strains...
January 15, 2017: Applied and Environmental Microbiology
https://www.readbyqxmd.com/read/27829029/inferring-gene-networks-for-strains-of-dehalococcoides-highlights-conserved-relationships-between-genes-encoding-core-catabolic-and-cell-wall-structural-proteins
#18
Cresten B Mansfeldt, Gretchen W Heavner, Annette R Rowe, Boris Hayete, Bruce W Church, Ruth E Richardson
The interpretation of high-throughput gene expression data for non-model microorganisms remains obscured because of the high fraction of hypothetical genes and the limited number of methods for the robust inference of gene networks. Therefore, to elucidate gene-gene and gene-condition linkages in the bioremediation-important genus Dehalococcoides, we applied a Bayesian inference strategy called Reverse Engineering/Forward Simulation (REFS™) on transcriptomic data collected from two organohalide-respiring communities containing different Dehalococcoides mccartyi strains: the Cornell University mixed community D2 and the commercially available KB-1® bioaugmentation culture...
2016: PloS One
https://www.readbyqxmd.com/read/27809491/1-2-dichloroethane-exposure-alters-the-population-structure-metabolism-and-kinetics-of-a-trichloroethene-dechlorinating-dehalococcoides-mccartyi-consortium
#19
Koshlan Mayer-Blackwell, Maeva Fincker, Olivia Molenda, Benjamin Callahan, Holly Sewell, Susan Holmes, Elizabeth A Edwards, Alfred M Spormann
Bioremediation of groundwater contaminated with chlorinated aliphatic hydrocarbons such as perchloroethene and trichloroethene can result in the accumulation of the undesirable intermediate vinyl chloride. Such accumulation can either be due to the absence of specific vinyl chloride respiring Dehalococcoides mccartyi or to the inhibition of such strains by the metabolism of other microorganisms. The fitness of vinyl chloride respiring Dehalococcoides mccartyi subpopulations is particularly uncertain in the presence of chloroethene/chloroethane cocontaminant mixtures, which are commonly found in contaminated groundwater...
November 15, 2016: Environmental Science & Technology
https://www.readbyqxmd.com/read/27800712/multi-template-homology-based-structural-model-of-l-2-haloacid-dehalogenase-dehl-from-rhizobium-sp-rc1
#20
Aliyu Adamu, Mohd Shahir Shamsir, Roswanira Abdul Wahab, Sepideh Parvizpour, Fahrul Huyop
Dehalogenases are of high interest due to their potential applications in bioremediation and in synthesis of various industrial products. DehL is an L-2-haloacid dehalogenase (EC 3.8.1.2) that catalyses the cleavage of halide ion from L-2-halocarboxylic acid to produce D-2-hydroxycarboxylic acid. Although DehL utilises the same substrates as the other L-2-haloacid dehalogenases, its deduced amino acid sequence is substantially different (<25%) from those of the rest L-2-haloacid dehalogenases. To date, the 3D structure of DehL is not available...
November 21, 2016: Journal of Biomolecular Structure & Dynamics
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