Lyashenko Andrey V, Zhukhlistova Nadegda E, Gabdoulkhakov Azat G, Zhukova Yuliya N, Voelter Wolfang, Zaitsev Viatcheslav N, Bento Isabel, Stepanova Elena V, Kachalova Galina S, Koroleva Ol'ga V, Cherkashyn Evgeniy A, Tishkov Vladimir I, Lamzin Victor S, Schirwitz Katja, Morgunova Ekaterina Yu, Betzel Christian, Lindley Peter F, Mikhailov Al'bert M
A. V. Shubnikov Institute of Crystallography, RAS, Leninskiy Prospect 59, 119333 Moscow, Russia.
Acta Crystallogr Sect F Struct Biol Cryst Commun. 2006 Oct 1;62(Pt 10):954-7. doi: 10.1107/S1744309106036578. Epub 2006 Sep 19.
Laccases are members of the blue multi-copper oxidase family that oxidize substrate molecules by accepting electrons at a mononuclear copper centre and transferring them to a trinuclear centre. Dioxygen binds to the trinuclear centre and, following the transfer of four electrons, is reduced to two molecules of water. Crystals of the laccase from Cerrena maxima have been obtained and X-ray data were collected to 1.9 A resolution using synchrotron radiation. A preliminary analysis shows that the enzyme has the typical laccase structure and several carbohydrate sites have been identified. The carbohydrate chains appear to be involved in stabilization of the intermolecular contacts in the crystal structure, thus promoting the formation of well ordered crystals of the enzyme. Here, the results of an X-ray crystallographic study on the laccase from the fungus Cerrena maxima are reported. Crystals that diffract well to a resolution of at least 1.9 A (R factor = 18.953%; R(free) = 23.835; r.m.s.d. bond lengths, 0.06 A; r.m.s.d. bond angles, 1.07 degrees) have been obtained despite the presence of glycan moieties. The overall spatial organization of C. maxima laccase and the structure of its copper-containing active centre have been determined by the molecular-replacement method using the laccase from Trametes versicolor (Piontek et al., 2002) as a structural template. In addition, four glycan-binding sites were identified and the 1.9 A X-ray data were used to determine the previously unknown primary structure of this protein. The identity (calculated from sequence alignment) between the C. maxima laccase and the T. versicolor laccase is about 87%. Tyr196 and Tyr372 show significant extra density at the ortho positions and this has been interpreted in terms of NO(2) substituents.
漆酶是蓝色多铜氧化酶家族的成员，通过在单核铜中心接受电子并将其转移到三核中心来氧化底物分子。双氧与三核中心结合，在四个电子转移后，被还原为两分子水。已获得来自大孢蜡蘑漆酶的晶体，并使用同步辐射将X射线数据收集至1.9 Å分辨率。初步分析表明该酶具有典型的漆酶结构，并已鉴定出几个碳水化合物位点。碳水化合物链似乎参与了晶体结构中分子间接触的稳定，从而促进了该酶有序晶体的形成。在此，报道了对真菌大孢蜡蘑漆酶的X射线晶体学研究结果。尽管存在聚糖部分，但已获得了能衍射至至少1.9 Å分辨率（R因子 = 18.953%；R（自由） = 23.835；均方根键长，0.06 Å；均方根键角，1.07度）的良好晶体。使用云芝漆酶（Piontek等人，2002年）作为结构模板，通过分子置换法确定了大孢蜡蘑漆酶的整体空间组织及其含铜活性中心的结构。此外，鉴定出四个聚糖结合位点，并利用1.9 Å的X射线数据确定了该蛋白质先前未知的一级结构。大孢蜡蘑漆酶与云芝漆酶之间的同一性（根据序列比对计算）约为87%。Tyr196和Tyr372在邻位显示出明显的额外密度，这已根据NO(2)取代基进行了解释。
