Structure Study of Vitamin B12 enzyme, Diol Dehydratase

得到了氰钴胺与催产克雷伯氏菌二醇脱水酶的两种晶型，并对其晶体结构进行了分析。根据相应的结晶条件，晶体属于两个不同的空间群。其中一种晶体属于P2_12_12_1空间群，晶胞尺寸为a=76.2，b=122.3，c=209.6Å，用同步辐射源的X射线束衍射率高达2.2Å。另一种晶体属于P21空间群，晶胞尺寸a=75.4，b=132.7，c=298.8Å，β=91.9°，衍射分辨率达3.0Å。形式I的三维结构是在2.2Å分辨率下确定的。二醇脱水酶以杂三聚体(αβγ)_2的二聚体形式存在。B_&lt；12&gt；分子以“碱基”方式结合在α和β亚基之间，即核苷酸部分的5，6-二甲基苯并咪唑与下轴位置的钴原子配位。α亚基包括一个(β/α)_8桶。底物和一种必需的钾离子深埋在桶内。基于钾离子与底物丙二醇的两个羟基之间独特的直接相互作用，提出了一种钾离子直接参与酶催化的方法。

Two crystal forms of Klebsiella oxytoca diol dehydratase complexed with cyanocobalamin have been obtained and crystallographic structure analysis have been performed. The crystals belong to two different space groups, according to the corresponding crystallization condition. One crystal (form I) belongs to the space group P2_12_12_1 with unit cell dimensions of a=76.2, b=122.3, c=209.6 Å, and diffracts up to 2.2Å resolution using an X-ray beam from a synchrotron radiation source. The other crystal (form II) belongs to the space group P2_1 with unit cell dimensions of a=75.4, b=132.7, c=298.8Å, β=91.9° and diffracts up to 3.0Å resolution. The three-dimensional structure of form I was determined at 2.2Å resolution. Diol dehydratase exists in a dimeric form of hetero-trimer, (αβγ)_2. The B_<12> molecule is bound between the α and β subunits in the "base-on" mode, that is, 5,6-dimethylbenzimidazole of the nucleotide moiety is coordinated to the cobalt atom in the lower axial position. The α subunit includes a (β/α)_8 barrel. The substrate and an essential potassium ion exist deeply buried inside the barrel. Based on unique direct interactions between potassium ion and two hydroxyl groups of the substrate, propanediol, direct participation of a potassium ion in enzyme catalysis is proposed.