Shibata N, Masuda J, Tobimatsu T, Toraya T, Suto K, Morimoto Y, Yasuoka N
Department of Life Science, Himeji Institute of Technology, Hyogo, Japan.
Structure. 1999 Aug 15;7(8):997-1008. doi: 10.1016/s0969-2126(99)80126-9.
Diol dehydratase is an enzyme that catalyzes the adenosylcobalamin (coenzyme B12) dependent conversion of 1,2-diols to the corresponding aldehydes. The reaction initiated by homolytic cleavage of the cobalt-carbon bond of the coenzyme proceeds by a radical mechanism. The enzyme is an alpha2beta2gamma2 heterooligomer and has an absolute requirement for a potassium ion for catalytic activity. The crystal structure analysis of a diol dehydratase-cyanocobalamin complex was carried out in order to help understand the mechanism of action of this enzyme.
The three-dimensional structure of diol dehydratase in complex with cyanocobalamin was determined at 2.2 A resolution. The enzyme exists as a dimer of heterotrimers (alphabetagamma)2. The cobalamin molecule is bound between the alpha and beta subunits in the 'base-on' mode, that is, 5,6-dimethylbenzimidazole of the nucleotide moiety coordinates to the cobalt atom in the lower axial position. The alpha subunit includes a (beta/alpha)8 barrel. The substrate, 1,2-propanediol, and an essential potassium ion are deeply buried inside the barrel. The two hydroxyl groups of the substrate coordinate directly to the potassium ion.
This is the first crystallographic indication of the 'base-on' mode of cobalamin binding. An unusually long cobalt-base bond seems to favor homolytic cleavage of the cobalt-carbon bond and therefore to favor radical enzyme catalysis. Reactive radical intermediates can be protected from side reactions by spatial isolation inside the barrel. On the basis of unique direct interactions between the potassium ion and the two hydroxyl groups of the substrate, direct participation of a potassium ion in enzyme catalysis is strongly suggested.
二醇脱水酶是一种催化依赖腺苷钴胺素（辅酶B12）将1,2 -二醇转化为相应醛类的酶。由辅酶钴 - 碳键的均裂引发的反应通过自由基机制进行。该酶是一种α2β2γ2异源寡聚体，催化活性绝对需要钾离子。为了帮助理解这种酶的作用机制，对二醇脱水酶 - 氰钴胺素复合物进行了晶体结构分析。
在2.2埃分辨率下确定了二醇脱水酶与氰钴胺素复合物的三维结构。该酶以异源三聚体的二聚体（αβγ）2形式存在。钴胺素分子以“碱基朝上”模式结合在α和β亚基之间，即核苷酸部分的5,6 - 二甲基苯并咪唑在较低轴向位置与钴原子配位。α亚基包括一个（β/α）8桶状结构。底物1,2 - 丙二醇和一个必需的钾离子深埋在桶状结构内部。底物的两个羟基直接与钾离子配位。
这是钴胺素结合“碱基朝上”模式的首个晶体学证据。异常长的钴 - 碱基键似乎有利于钴 - 碳键的均裂，因此有利于自由基酶催化。反应性自由基中间体可以通过桶状结构内部的空间隔离来避免副反应。基于钾离子与底物的两个羟基之间独特的直接相互作用，强烈表明钾离子直接参与酶催化。
