Mancia F, Keep N H, Nakagawa A, Leadlay P F, McSweeney S, Rasmussen B, Bösecke P, Diat O, Evans P R
MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, UK.
Structure. 1996 Mar 15;4(3):339-50. doi: 10.1016/s0969-2126(96)00037-8.
The enzyme methylmalonyl-coenzyme A (CoA) mutase, an alphabeta heterodimer of 150 kDa, is a member of a class of enzymes that uses coenzyme B12 (adenosylcobalamin) as a cofactor. The enzyme induces the formation of an adenosyl radical from the cofactor. This radical then initiates a free-radical rearrangement of its substrate, succinyl-CoA, to methylmalonyl-CoA.
Reported here is the crystal structure at 2 A resolution of methylmalonyl-CoA mutase from Propionibacterium shermanii in complex with coenzyme B12 and with the partial substrate desulpho-CoA (lacking the succinyl group and the sulphur atom of the substrate). The coenzyme is bound by a domain which shares a similar fold to those of flavodoxin and the B12-binding domain of methylcobalamin-dependent methionine synthase. The cobalt atom is coordinated, via a long bond, to a histidine from the protein. The partial substrate is bound along the axis of a (beta/alpha)8 TIM barrel domain.
The histidine-cobalt distance is very long (2.5 A compared with 1.95-2.2 A in free cobalamins), suggesting that the enzyme positions the histidine in order to weaken the metal-carbon bond of the cofactor and favour the formation of the initial radical species. The active site is deeply buried, and the only access to it is through a narrow tunnel along the axis of the TIM barrel domain.
甲基丙二酰辅酶A变位酶是一种150 kDa的αβ异二聚体酶，属于一类以辅酶B12（腺苷钴胺素）为辅因子的酶。该酶从辅因子诱导形成腺苷自由基。然后这个自由基引发其底物琥珀酰辅酶A的自由基重排，生成甲基丙二酰辅酶A。
本文报道了来自谢氏丙酸杆菌的甲基丙二酰辅酶A变位酶与辅酶B12及部分底物脱硫辅酶A（缺乏底物的琥珀酰基团和硫原子）复合物在2 Å分辨率下的晶体结构。辅酶由一个结构域结合，该结构域与黄素氧还蛋白和钴胺素依赖性甲硫氨酸合酶的B12结合结构域具有相似的折叠方式。钴原子通过一个长键与蛋白质中的一个组氨酸配位。部分底物沿着一个（β/α）8 TIM桶状结构域的轴结合。
组氨酸 - 钴的距离非常长（2.5 Å，而游离钴胺素中的距离为1.95 - 2.2 Å），这表明该酶将组氨酸定位以削弱辅因子的金属 - 碳键并有利于初始自由基物种 的形成。活性位点深埋，唯一通向它的途径是通过沿着TIM桶状结构域轴的一条狭窄通道。
