Crystallization and initial X-ray analysis of phenoxazinone synthase from Streptomyces antibioticus.
Phenoxazinone synthase, an oligomeric multicopper oxidase produced by Streptomyces antibioticus, is responsible for the six-electron oxidative coupling of two molecules of 4-methyl 3-hydroxyanthraniloyl pentapeptide to form the phenoxazinone chromophore of the antineoplastic agent actinomycin D. Spectroscopic studies have shown that the enzyme contains one type I (blue) and three to four type II copper centers. However, the exact arrangement of the copper centers in this multicopper oxidase is unknown. As a first step towards determining the three-dimensional structure of the enzyme, phenoxazinone synthase has been crystallized. The hexameric form of phenoxazinone synthase was purified from 72 h cultures of S. lividans containing the plasmid pIJ702. Purified hexamers were concentrated to 75 mg ml(-1) and used to grow two forms of crystals. Data collected from the two crystal forms were processed in two separate space groups. Crystals of both forms were grown at 288 K using the sitting-drop vapour-diffusion method. Native data sets extending to resolutions of 3.35 and 2.30 A have been collected and processed in space groups R32 and P1, respectively.
Department of Chemistry and Biochemistry, Arizona State University, Tempe 85287-1604, USA.
Alex W
AW
Smith
Ana
A
Camara-Artigas
Charles
C
Olea
Wilson A
WA
Francisco
James P
JP
Allen
D003460:Crystallization; D018360:Crystallography, X-Ray; D010088:Oxidoreductases / Q000737:chemistry*; D013303:Streptomyces antibioticus / Q000201:enzymology*
D010088:Oxidoreductases; C023944:2-aminophenol oxidase