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Updated: Jul 20, 2026
X-Ray Crystallography to Study the Oligomeric State Transition of the Thermotoga maritima M42 Aminopeptidase TmPep1050
Published on: May 13, 2020
The purification, crystallization and preliminary structural characterization of PhzM, a phenazine-modifying
Neelakshi Gohain1, Linda S Thomashow, Dmitri V Mavrodi
1Max-Planck-Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund, Germany.
Abstract:
Pyocyanin, phenazine-1-carboxylic acid and more than 70 related compounds collectively known as phenazines are produced by various species of Pseudomonas, including the fluorescent pseudomonad P. aeruginosa, a Gram-negative opportunistic pathogen in humans and animals. P. aeruginosa synthesizes a characteristic blue water-soluble compound called pyocyanin (1-hydroxy-5-methyl-phenazine). Two enzymes designated PhzM and PhzS are involved in the terminal steps of its synthesis and very little is known about these enzymes. In this study, PhzM, a dimeric S-adenosylmethionine-dependent methyltransferase, was purified and crystallized from PEG 3350/sodium cacodylate/sodium citrate pH 6.5. The crystals belong to space group P1, with unit-cell parameters a = 46.1, b = 61.8, c = 69.6 A, alpha = 96.3, beta = 106.6, gamma = 106.9 degrees . They contain one dimer in the asymmetric unit and diffract to a resolution of 1.8 A. Anomalous data to 2.3 A resolution have been collected from seleno-L-methionine-labelled PhzM.
Insights
Researchers purified and crystallized the PhzM enzyme, crucial for synthesizing pyocyanin in Pseudomonas aeruginosa. This structural study provides insights into the enzyme
Area of Science:
- Microbiology and Biochemistry
- Structural Biology
Background:
- Phenazines, including pyocyanin, are secondary metabolites produced by Pseudomonas species.
- Pseudomonas aeruginosa, an opportunistic pathogen, synthesizes pyocyanin, a blue pigment.
- Enzymes PhzM and PhzS are involved in the final stages of pyocyanin biosynthesis, but their structures are largely unknown.
Purpose of the Study:
- To elucidate the structure of the PhzM enzyme, an S-adenosylmethionine-dependent methyltransferase.
- To understand the structural basis for pyocyanin synthesis in Pseudomonas aeruginosa.
Main Methods:
- Purification of the dimeric PhzM enzyme.
- Crystallization of PhzM using PEG 3350/sodium cacodylate/sodium citrate buffer at pH 6.5.
- X-ray diffraction analysis of native and seleno-L-methionine-labeled PhzM crystals.
- Determination of unit-cell parameters and space group (P1).
Main Results:
- PhzM crystals diffracting to 1.8 A resolution were obtained.
- The crystal structure belongs to space group P1 with specific unit-cell parameters.
- The asymmetric unit contains one PhzM dimer.
- Anomalous diffraction data were collected to 2.3 A resolution from labeled crystals.
Conclusions:
- The study presents the first structural characterization of the PhzM enzyme.
- The obtained crystal structure provides a foundation for understanding the mechanism of pyocyanin biosynthesis.
- Further structural studies will aid in understanding phenazine production in Pseudomonas species.
