Matera I, Ferraroni M, Bürger S, Stolz A, Briganti F
Department of Chemistry, University of Florence, Via della Lastruccia 3, 50019 Sesto Fiorentino, Florence, Italy.
Acta Crystallogr Sect F Struct Biol Cryst Commun. 2006 Jun 1;62(Pt 6):553-5. doi: 10.1107/S1744309106016435. Epub 2006 May 31.
Salicylate 1,2-dioxygenase, a new ring-fission dioxygenase from the naphthalenesulfonate-degrading strain Pseudaminobacter salicylatoxidans which oxidizes salicylate to 2-oxohepta-3,5-dienedioic acid by a novel ring-fission mechanism, has been crystallized. Diffraction-quality crystals of salicylate 1,2-dioxygenase were obtained using the sitting-drop vapour-diffusion method at 277 K from a solution containing 10%(w/v) ethanol, 6%(w/v) PEG 400, 0.1 M sodium acetate pH 4.6. Crystals belong to the primitive tetragonal space group P4(3)2(1)2 or P4(1)2(1)2, with unit-cell parameters a = 133.3, c = 191.51 A. A complete data set at 100 K extending to a maximum resolution of 2.9 A was collected at a wavelength of 0.8423 A. Molecular replacement using the coordinates of known extradiol dioxygenases structures as a model has so far failed to provide a solution for salicylate 1,2-dioxygenase. Attempts are currently being made to solve the structure of the enzyme by MAD experiments using the anomalous signal of the catalytic Fe(II) ions. The salicylate 1,2-dioxygenase structural model will assist in the elucidation of the catalytic mechanism of this ring-fission dioxygenase from P. salicylatoxidans, which differs markedly from the known gentisate 1,2-dioxygenases or 1-hydroxy-2-naphthoate dioxygenases because of its unique ability to oxidatively cleave salicylate, gentisate and 1-hydroxy-2-naphthoate with high catalytic efficiency.
水杨酸1,2-双加氧酶是一种来自降解萘磺酸盐的菌株水杨酸盐氧化假氨基杆菌的新型开环双加氧酶，它通过一种新颖的开环机制将水杨酸氧化为2-氧代庚-3,5-二烯二酸，目前已结晶。使用坐滴气相扩散法，在277 K下从含有10%（w/v）乙醇、6%（w/v）聚乙二醇400、0.1 M pH 4.6醋酸钠的溶液中获得了衍射质量良好的水杨酸1,2-双加氧酶晶体。晶体属于原始四方空间群P4(3)2(1)2或P4(1)2(1)2，晶胞参数a = 133.3，c = 191.51 Å。在100 K下，以0.8423 Å的波长收集了一个完整的数据集，最大分辨率为2.9 Å。到目前为止，以已知的二醇双加氧酶结构坐标为模型进行分子置换未能为水杨酸1,2-双加氧酶提供解决方案。目前正在尝试通过利用催化Fe(II)离子的异常信号进行MAD实验来解析该酶的结构。水杨酸1,2-双加氧酶的结构模型将有助于阐明来自水杨酸盐氧化假氨基杆菌的这种开环双加氧酶的催化机制，该机制与已知的龙胆酸1,2-双加氧酶或1-羟基-2-萘甲酸双加氧酶明显不同，因为它具有以高催化效率氧化裂解水杨酸、龙胆酸和1-羟基-2-萘甲酸的独特能力。
