Crystallographic evidence that the dinuclear copper center of tyrosinase is flexible during catalysis
Crystallographic evidence that the dinuclear copper center of tyrosinase is flexible during catalysis
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DOI：
10.1074/jbc.m509785200
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发表时间：
2006-03-31
影响因子：
4.8
通讯作者：
Sugiyama, M
中科院分区：
文献类型：
--
作者：
Matoba, Y;Kumagai, T;Sugiyama, M
在高分辨率下，我们测定了与ORF378的络合物中铜结合和无金属酪氨酸酶的晶体结构，因为它帮助两个铜(II)离子进入酪氨酸酶催化中心。这些结构表明，球蛋白覆盖在酪氨酸酶的疏水分子表面，并干扰底物酪氨酸与酪氨酸酶催化部位的结合。球茎蛋白由一个六链的β-折叠和一个α-螺旋组成，与蛋白质数据库中存储的所有蛋白质没有相似性。虽然酪氨酸酶和邻苯二酚氧化酶属于3型铜蛋白家族，但后者缺乏单加氧酶活性。催化活性的差异是基于结构观察，即酪氨酸酶活性中心上方存在一个大的空位，两个铜离子的六个His配体中的一个是高度灵活的。酪氨酸酶的这些结构特征表明，在催化单酚邻位羟基化的反应中，两个铜(II)离子中的一个被底物上的过氧化氢生成的氧配位。我们的结晶学研究表明，双核铜形成的酪氨酸酶活性中心在催化过程中是柔性的。
At high resolution, we determined the crystal structures of copper-bound and metal-free tyrosinase in a complex with ORF378 designated as a "caddie" protein because it assists with transportation of two Cu(II) ions into the tyrosinase catalytic center. These structures suggest that the caddie protein covers the hydrophobic molecular surface of tyrosinase and interferes with the binding of a substrate tyrosine to the catalytic site of tyrosinase. The caddie protein, which consists of one six-stranded beta-sheet and one alpha-helix, has no similarity with all proteins deposited into the Protein Data Bank. Although tyrosinase and catechol oxidase are classified into the type 3 copper protein family, the latter enzyme lacks monooxygenase activity. The difference in catalytic activity is based on the structural observations that a large vacant space is present just above the active center of tyrosinase and that one of the six His ligands for the two copper ions is highly flexible. These structural characteristics of tyrosinase suggest that, in the reaction that catalyzes the ortho-hydroxylation of monophenol, one of the two Cu(II) ions is coordinated by the peroxide-originated oxygen bound to the substrate. Our crystallographic study shows evidence that the tyrosinase active center formed by dinuclear coppers is flexible during catalysis.