ID 4 GLUTAMYL RADICAL AS INTERMEDIATE IN CARBON SKELETON REARRANGEMENT

使用一系列2H和13C标记的谷氨酸盐作为底物 对于依赖辅酶B12的谷氨酸变位酶，它平衡 (S)-谷氨酸与(2S，3S)-3-甲基天冬氨酸。这些化合物 含有碳链的C-2、C-3或C-4的同位素： [2-2H]，[3-2H2]，[4-2H2]，[2，3，3，4，4-2H5]，[2-13C]，[3-13C]，和 [4-13C]谷氨酸。每个反应都通过EPR波谱进行监测 发现了类似的信号，其特征为g‘xy=2.1g’z=1.985 A‘=5.0mT。对光谱数据的解释是由 模拟结果与实验结果吻合较好。这种方法 支撑了自由基对形成的想法，包括 CoB(II)丙胺与有机自由基间的相互作用 6.6+/-0.9A。观察到的超精细耦合的比较 未标记的谷氨酸与来自标记的谷氨酸盐的谷氨酸启用 自由基对的主要贡献者被确定为 4-甘氨酰基。这些发现支持目前公认的 谷氨酸变位酶反应的机理，即该过程是 通过从谷氨酸的C-4中提取氢原子引发 脱氧腺苷5‘-脱氧腺苷自由基，通过均相裂解产生 辅酶B21上的辅酶C-Sigma键。

A series of 2H- and 13C-labeled glutamates were used as substrates for coenzyme B12-dependent glutamate mutase, which equilibrates (S)-glutamate with (2S, 3S)-3-methylasparate. These compounds contained the isotopes at C-2, C-3 or C-4 of the carbon chain: [2-2H], [3-2H2], [4-2H2], [2,3,3,4,4-2H5], [2-13C], [3-13C], and [4-13C]glutamate. Each reaction was monitored by EPR spectroscopy and revealed a similar signal characterized by g'xy=2.1,g'z=1.985 and A'=5.0mT. The interpretation of the spectral data was aided by simulations which gave close agreement with experiment. This approach underpinned the idea of the formation of a radical pair, consisting of cob(II)alamin interacting with an organic radical at a distance of 6.6+/- 0.9A. Comparison of the hyperfine couplings observed with unlabeled glutamate with those from the labeled glutamates enabled a principle contributor to the radical pair to be identiied as the 4-glytamyl radical. These findings support the currently accepted mechanism for the glutamate mutase reaction, i.e. the process is initiated through hydrogen atom abstraction from C-4 of glutamate by the 5'-deoxyadenosyl radical, which is derived by homolysis of the Co-C sigma-bond o the coenzyme B21.