Interactions of diol dehydrase and 3',4'-anhydroadenosylcobalamin: suicide inactivation by electron transfer.
Interactions of diol dehydrase and 3',4'-anhydroadenosylcobalamin: suicide inactivation by electron transfer.
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二醇脱水酶和 3,4-脱水腺苷钴胺素的相互作用：通过电子转移自杀失活。
DOI：
10.1021/bi011947w
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发表时间：
2002
期刊：
影响因子：
2.9
通讯作者：
Frey,PerryA
中科院分区：
文献类型：
--
作者：
Magnusson,OlafurTh;Frey,PerryA
3 ',4 ' -无氢腺苷钴胺素（anAdoCbl）是腺苷钴胺素（AdoCbl）辅酶的类似物。弗雷，P. A. (2000)J。点。化学。Soc. 122, 8807−8813)。该化合物支持二醇脱氢酶的活性为AdoCbl的0.02%。然而，在副反应中，adocbl通过电子转移机制诱导自杀失活。用分光光度法检测cob(II)alamin，观察到在二醇脱水酶活性位点，anadocbl的Co−C键发生了均匀的裂解。酶结合的cob(II)alamin向cob(III)alamin的厌氧转化，在无底物和存在底物的情况下，表明辅酶衍生的5 ' -脱氧-3 ',4 ' -无氢腺苷-5 ' -酰基起氧化剂的作用。通过高效液相色谱法、质谱法和核磁共振波谱法确定，3 ',5 ' -二脱氧腺苷-4 ',5 ' -烯作为核苷裂解产物的化学计量形成支持了这一假设。在氧化氘中进行的实验表明，产物中加入了一个溶剂交换质子。这些数据与cob(II)alamin在一个电子转移到烯丙基5 ' -脱氧-3 ',4 ' -无氢腺苷自由基后形成的瞬时烯丙基阴离子的中间形成一致。通过对纯化产物的光谱表征，证实了C3 '位置的选择性质子化作用。本研究提供了一个由自由基中间体类似物的副反应引起的自由基酶自杀性失活的例子。
3‘,4‘-Anhydroadenosylcobalamin (anAdoCbl) is an analogue of the adenosylcobalamin (AdoCbl) coenzyme (Magnusson, O.Th., and Frey, P. A. (2000)J. Am. Chem. Soc. 122, 8807−8813). This compound supports activity for diol dehydrase at 0.02% of that observed with AdoCbl. In a side reaction, however,anAdoCbl induces suicide inactivation by an electron-transfer mechanism. Homolytic cleavage of the Co−C bond ofanAdoCbl at the active site of diol dehydrase was observed by spectrophotometric detection of cob(II)alamin. Anaerobic conversion of enzyme bound cob(II)alamin to cob(III)alamin, both in the absence and presence of substrate, indicates that the coenzyme derived 5‘-deoxy-3‘,4‘-anhydroadenosine-5‘-yl serves as the oxidizing agent. This hypothesis is supported by the stoichiometric formation of 3‘,5‘-dideoxyadenosine-4‘,5‘-ene as the nucleoside cleavage product, as determined by high-performance liquid chromatography, mass spectrometry, and nuclear magnetic resonance spectroscopy. Experiments performed in deuterium oxide show that a single solvent exchangeable proton is incorporated into the product. These data are consistent with the intermediate formation of a transient allylic anion formed after one electron transfer from cob(II)alamin to the allylic 5‘-deoxy-3‘,4‘-anhydroadenosyl radical. Selective protonation at C3‘ was demonstrated by spectroscopic characterization of the purified product. This study provides an example of suicide inactivation of a radical enzyme brought about by a side reaction of an analogue of the radical intermediate.