Q51135663
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We report the biochemical characterization of a novel haloalkane dehalogenase, DatA, isolated from the plant pathogen Agrobacterium tumefaciens C58. DatA possesses a peculiar pair of halide-stabilizing residues, Asn-Tyr, which have not been reported to play this role in other known haloalkane dehalogenases. DatA has a number of other unique characteristics, including substrate-dependent and cooperative kinetics, a dimeric structure, and excellent enantioselectivity toward racemic mixtures of chiral brominated alkanes and esters.
- MeSH
- Agrobacterium tumefaciens enzymologie genetika metabolismus MeSH
- alkany metabolismus MeSH
- DNA bakterií chemie genetika MeSH
- estery metabolismus MeSH
- hydrolasy genetika izolace a purifikace metabolismus MeSH
- molekulární sekvence - údaje MeSH
- multimerizace proteinu MeSH
- rostliny mikrobiologie MeSH
- sekvenční analýza DNA MeSH
- stereoizomerie MeSH
- substrátová specifita MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Capsidiol is a bicyclic sesquiterpene, which accumulates extracellularly in plants, and has been isolated from many types of Solanaceae. It acts as a phytoalexin produced by Nicotiana tabacum in response to pathogens. Capsidiol has antifungal activity and is formed first in tobacco and pepper plants after infestation. The amount of capsidiol in tobacco cell suspension culture has been previously determined by solid-phase extraction and organic solvent extraction with thin-layer chromatography or gas chromatography analysis. A high-performance liquid chromatography method with UV detection at 210 nm on a C(8) column utilizing both extraction methods was developed to analyze capsidiol in suspension cell culture. The HPLC method was linear in the concentration range of 0.1-2.0 mg/L. The lower limit of quantitation was 0.1 mg/L. Organic solvent extraction and solid-phase extraction methods were compared. Both methods are generally similar in their overall efficiency (82% and 75%, respectively), but eliminations of interfering compounds are different. The relative standard deviation across five extractions of known amounts of capsidiol from plant sample was less than 5.1%. The relative standard deviation across five elicitations of cell cultures was less than 5.9%. Gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry analysis of capsidiol was performed, and corresponding mass spectra are presented.
- MeSH
- chemická frakcionace MeSH
- chromatografie kapalinová MeSH
- hmotnostní spektrometrie MeSH
- kultivované buňky MeSH
- lineární modely MeSH
- plynová chromatografie s hmotnostně spektrometrickou detekcí MeSH
- reprodukovatelnost výsledků MeSH
- rostlinné extrakty analýza MeSH
- seskviterpeny analýza MeSH
- tabák chemie MeSH
- vysokoúčinná kapalinová chromatografie metody MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
An enzyme-based biosensor was developed by co-immobilization of purified enzyme haloalkane dehalogenase (EC 3.8.1.5) and a fluorescence pH indicator on the tip of an optical fiber. Haloalkane dehalogenase catalyzes hydrolytic dehalogenation of halogenated aliphatic hydrocarbons, which is accompanied by a pH change influencing the fluorescence of the indicator. The pH sensitivity of several fluorescent dyes was evaluated. The selected indicator 5(6)-carboxyfluorescein was conjugated with bovine serum albumin and its reaction was tested under different immobilization conditions. The biosensor was prepared by cross-linking of the conjugate in tandem with haloalkane dehalogenase using glutaraldehyde vapor. The biosensor, stored for 24 h in 50 mM phosphate buffer (pH 7.5) prior to measurement, was used after 15 min of equilibration, the halogenated compound was added, and the response was monitored for 30 min. Calibration of the biosensor with 1,2-dibromoethane and 3-chloro-2-(chloromethyl)-1-propene showed an excellent linear dependence, with detection limits of 0.133 and 0.014 mM, respectively. This biosensor provides a new tool for continuous in situ monitoring of halogenated environmental pollutants.
- MeSH
- biosenzitivní techniky přístrojové vybavení metody MeSH
- enzymy imobilizované chemie MeSH
- fluorescenční barviva chemie MeSH
- halogenované uhlovodíky analýza chemie MeSH
- molekulární struktura MeSH
- skot MeSH
- technologie optických vláken MeSH
- zvířata MeSH
- Check Tag
- skot MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Chemical warfare agents are toxic compounds showing negative effects on living organisms. This review focuses on detection of nerve and blister agents, which are two most important classes of chemical warfare agents. Main attention is paid to the construction and use of biosensors. Acetylcholinesterase, butyrylcholinesterase and phosphotriesterase are presented as convenient biorecognition components of biosensors for detection of nerve agents, while dehalogenases are useful for detection of mustard agents.
Chirality is a phenomenon that pervades the life on the Earth and has a tremendous importance for our daily lives. Many pharmaceuticals, agrochemicals, food additives, pheromones, fragrances and cosmetics are chiral. Chiral compounds play an essential role in biological systems, mainly for chemical communication among living organisms, serving as sex pheromones, aggregation pheromones, alarm pheromones, trail pheromones, attractants or repellent agents. This review summarizes our current knowledge of the biologically relevant chiral aliphatic hydroxy compounds, which are divided into eight classes according to their chemical structure: primary alcohols, secondary alcohols, tertiary alcohols, glycols and diols, hydroxy ketones, hydroxy carboxylic acids, hydroxy carboxylic esters and hydroxy amines. Information on biological functions and practical applications is summarized for each class. This information could be of interest to chemists, biochemists, biologists and pharmacists. A review with 475 references.
- MeSH
- biologické přípravky MeSH
- stereoizomerie MeSH
- Publikační typ
- přehledy MeSH
