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Title: The purification, crystallization and preliminary structural characterization of FAD-dependent monooxygenase PhzS, a phenazine-modifying enzyme from Pseudomonas aeruginosa

Abstract

PhzS, an FAD-dependent monooxygenase that catalyzes a reaction involved in the biosynthesis of the virulence factor pyocyanin in P. aeruginosa, was cloned, overexpressed and crystallized. Data collection from native and seleno-l-methionine-labelled crystals is reported. The blue chloroform-soluble bacterial metabolite pyocyanin (1-hydroxy-5-methyl-phenazine) contributes to the survival and virulence of Pseudomonas aeruginosa, an important Gram-negative opportunistic pathogen of humans and animals. Little is known about the two enzymes, designated PhzM and PhzS, that function in the synthesis of pyocyanin from phenazine-1-carboxylic acid. In this study, the FAD-dependent monooxygenase PhzS was purified and crystallized from lithium sulfate/ammonium sulfate/sodium citrate pH 5.5. Native crystals belong to space group C2, with unit-cell parameters a = 144.2, b = 96.2, c = 71.7 Å, α = γ = 90, β = 110.5°. They contain two monomers of PhzS in the asymmetric unit and diffract to a resolution of 2.4 Å. Seleno-l-methionine-labelled PhzS also crystallizes in space group C2, but the unit-cell parameters change to a = 70.6, b = 76.2, c = 80.2 Å, α = γ = 90, β = 110.5° and the diffraction limit is 2.7 Å.

Authors:
 [1];  [2];  [2];  [1]
  1. Max-Planck-Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund (Germany)
  2. Department of Plant Pathology, Washington State University, Pullman, Washington 99164-6430 (United States)
Publication Date:
OSTI Identifier:
22356362
Resource Type:
Journal Article
Journal Name:
Acta Crystallographica. Section F
Additional Journal Information:
Journal Volume: 62; Journal Issue: Pt 10; Other Information: PMCID: PMC2225181; PMID: 17012792; PUBLISHER-ID: fw5101; OAI: oai:pubmedcentral.nih.gov:2225181; Copyright (c) International Union of Crystallography 2006; Country of input: International Atomic Energy Agency (IAEA); Journal ID: ISSN 1744-3091
Country of Publication:
United Kingdom
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; AMMONIUM SULFATES; CHLOROFORM; CITRATES; CRYSTALLIZATION; CRYSTALS; DIFFRACTION; MONOMERS; OXYGEN; RESOLUTION; SODIUM COMPOUNDS; SPACE GROUPS

Citation Formats

Gohain, Neelakshi, Thomashow, Linda S., USDA Agricultural Research Service, Root Disease and Biological Control Research Unit, Pullman, Washington 99164-6430, Mavrodi, Dmitri V., and Blankenfeldt, Wulf. The purification, crystallization and preliminary structural characterization of FAD-dependent monooxygenase PhzS, a phenazine-modifying enzyme from Pseudomonas aeruginosa. United Kingdom: N. p., 2006. Web. doi:10.1107/S1744309106034464.
Gohain, Neelakshi, Thomashow, Linda S., USDA Agricultural Research Service, Root Disease and Biological Control Research Unit, Pullman, Washington 99164-6430, Mavrodi, Dmitri V., & Blankenfeldt, Wulf. The purification, crystallization and preliminary structural characterization of FAD-dependent monooxygenase PhzS, a phenazine-modifying enzyme from Pseudomonas aeruginosa. United Kingdom. https://doi.org/10.1107/S1744309106034464
Gohain, Neelakshi, Thomashow, Linda S., USDA Agricultural Research Service, Root Disease and Biological Control Research Unit, Pullman, Washington 99164-6430, Mavrodi, Dmitri V., and Blankenfeldt, Wulf. 2006. "The purification, crystallization and preliminary structural characterization of FAD-dependent monooxygenase PhzS, a phenazine-modifying enzyme from Pseudomonas aeruginosa". United Kingdom. https://doi.org/10.1107/S1744309106034464.
@article{osti_22356362,
title = {The purification, crystallization and preliminary structural characterization of FAD-dependent monooxygenase PhzS, a phenazine-modifying enzyme from Pseudomonas aeruginosa},
author = {Gohain, Neelakshi and Thomashow, Linda S. and USDA Agricultural Research Service, Root Disease and Biological Control Research Unit, Pullman, Washington 99164-6430 and Mavrodi, Dmitri V. and Blankenfeldt, Wulf},
abstractNote = {PhzS, an FAD-dependent monooxygenase that catalyzes a reaction involved in the biosynthesis of the virulence factor pyocyanin in P. aeruginosa, was cloned, overexpressed and crystallized. Data collection from native and seleno-l-methionine-labelled crystals is reported. The blue chloroform-soluble bacterial metabolite pyocyanin (1-hydroxy-5-methyl-phenazine) contributes to the survival and virulence of Pseudomonas aeruginosa, an important Gram-negative opportunistic pathogen of humans and animals. Little is known about the two enzymes, designated PhzM and PhzS, that function in the synthesis of pyocyanin from phenazine-1-carboxylic acid. In this study, the FAD-dependent monooxygenase PhzS was purified and crystallized from lithium sulfate/ammonium sulfate/sodium citrate pH 5.5. Native crystals belong to space group C2, with unit-cell parameters a = 144.2, b = 96.2, c = 71.7 Å, α = γ = 90, β = 110.5°. They contain two monomers of PhzS in the asymmetric unit and diffract to a resolution of 2.4 Å. Seleno-l-methionine-labelled PhzS also crystallizes in space group C2, but the unit-cell parameters change to a = 70.6, b = 76.2, c = 80.2 Å, α = γ = 90, β = 110.5° and the diffraction limit is 2.7 Å.},
doi = {10.1107/S1744309106034464},
url = {https://www.osti.gov/biblio/22356362}, journal = {Acta Crystallographica. Section F},
issn = {1744-3091},
number = Pt 10,
volume = 62,
place = {United Kingdom},
year = {Sun Oct 01 00:00:00 EDT 2006},
month = {Sun Oct 01 00:00:00 EDT 2006}
}