4-Chlorocatechol 1,2-dioxygenase from the chlorophenol-utilizing Gram-positive Rhodococcus opacus 1CP: crystallization and preliminary crystallographic analysis.
4-Chlorocatechol 1,2-dioxygenase (4-ClC1,2DO) from the Gram-positive bacterium Rhodococcus opacus (erythropolis) 1CP, an enzyme involved in the aerobic biodegradation of chloroaromatic compounds, has been crystallized. 4-ClC1,2DO, which specifically catalyzes the intradiol cleavage of 4-substituted catechols, which are intermediates in the degradation of a variety of aromatic pollutants, to the corresponding maleylacetates, has recently been purified to homogeneity. The enzyme is an homodimer composed of two identical subunits in an alpha(2)-type quaternary structure; it has a molecular weight of about 29 kDa per monomer and contains one Fe(III) and one Mn(II) ion per homodimer. Hexagonal crystals grown in 1.6 M ammonium sulfate, 0.1 M sodium chloride, 100 mM Tris-HCl pH 9.0, 5-15% glycerol were successfully frozen under liquid nitrogen, adding 30% glycerol to the mother-liquor solution as a cryoprotectant. A complete data set was collected at 2.8 A resolution using the EMBL beamline BW7A at the DORIS storage ring, DESY, Hamburg, Germany with a MAR CCD detector and a wavelength of 1.01 A. The crystals belong to the primitive hexagonal space group P6(3)22, with unit-cell parameters a = 90.4, c = 307.5 A. This is the first intradiol dioxygenase which specifically catalyzes the cleavage of chlorocatechols in Gram-positive bacteria to give diffraction-quality crystals.
Ferraroni
Marta
M
Dipartimento di Chimica, Università di Firenze, Via della Lastruccia 3, I-50019 Sesto Fiorentino (FI), Italy.
Ruiz Tarifa
Maria Yolanda
MY
Briganti
Fabrizio
F
Scozzafava
Andrea
A
Mangani
Stefano
S
Solyanikova
Inna P
IP
Kolomytseva
Marina P
MP
Golovleva
Ludmilla
L
D002733:Chlorophenols / Q000378:metabolism; D003460:Crystallization; D018360:Crystallography, X-Ray; D049308:Dioxygenases; D010105:Oxygenases / Q000737:chemistry*; D011487:Protein Conformation; D012240:Rhodococcus / Q000201:enzymology*
D002733:Chlorophenols; D010105:Oxygenases; D049308:Dioxygenases; C053101:catechol oxygenase II