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Crystallization and preliminary X-ray studies of methyl parathion hydrolase from Pseudomonas sp. WBC-3
Lei Sun1, Yanjie Dong, Yafeng Zhou
1National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Science and Laboratory of Structural Biology, Tsinghua University, Beijing, People's Republic of China.
Abstract:
Methyl parathion hydrolase (MPH) from Pseudomonas sp. WBC-3, an enzyme that catalyzes the degradation of methyl parathion (O,O-dimethyl O-p-nitrophenyl phosphorothioate; MP), has been purified and crystallized by the hanging-drop vapour-diffusion method. The crystals were grown at 291 K using a precipitant solution consisting of 30% PEG 400, 0.1 M sodium acetate pH 4.6, 0.1 M CdCl(2). MPH is a zinc-containing enzyme judged by inductively coupled plasma mass-spectrometric (ICP-MS) analysis. Multiple-wavelength anomalous dispersive X-ray data were collected at 2.5 A resolution from a single crystal on beamline 41XU at SPring-8. The crystal belongs to space group P4(3)2(1)2, with unit-cell parameters a = 84.94, b = 84.94, c = 200.38 A, alpha = beta = gamma = 90 degrees. The asymmetric unit contains two molecules and has a solvent content of approximately 52%. Crystal structure determination is in progress.
Insights
Methyl parathion hydrolase (MPH) from Pseudomonas sp. WBC-3 was purified and crystallized. This enzyme aids in methyl parathion degradation and its crystal structure determination is underway.
Area of Science:
- Biochemistry
- Enzymology
- Structural Biology
Background:
- Methyl parathion is a toxic organophosphate pesticide.
- Methyl parathion hydrolase (MPH) from Pseudomonas sp. WBC-3 effectively degrades methyl parathion.
- Understanding MPH structure is crucial for developing detoxification strategies.
Purpose of the Study:
- To purify and crystallize Methyl parathion hydrolase (MPH) from Pseudomonas sp. WBC-3.
- To initiate the structural determination of MPH.
- To investigate the biochemical properties of MPH.
Main Methods:
- Enzyme purification and crystallization using the hanging-drop vapour-diffusion method.
- Inductively coupled plasma mass-spectrometric (ICP-MS) analysis for metal content.
- X-ray diffraction data collection at 2.5 Å resolution.
Main Results:
- Purified and crystallized MPH from Pseudomonas sp. WBC-3.
- MPH is a zinc-containing enzyme.
- Crystal data: Space group P4(3)2(1)2, unit-cell parameters a = 84.94, b = 84.94, c = 200.38 Å.
- The asymmetric unit contains two molecules with ~52% solvent content.
Conclusions:
- Successful crystallization of MPH provides a foundation for structure determination.
- The presence of zinc suggests its importance in the enzyme's catalytic mechanism.
- Further structural analysis will elucidate MPH's mechanism in methyl parathion degradation.
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