Paramagnetic centers in the nickel-containing, deazaflavin-reducing hydrogenase from Methanobacterium thermoautotrophicum.
Paramagnetic centers in the nickel-containing, deazaflavin-reducing hydrogenase from Methanobacterium thermoautotrophicum.
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来自热自养甲烷杆菌的含镍脱氮黄素还原氢化酶中的顺磁性中心。
DOI：
10.1073/pnas.80.2.378
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发表时间：
1983
影响因子：
11.1
通讯作者：
Walsh,C
中科院分区：
文献类型：
--
作者：
Kojima,N;Fox,JA;Hausinger,RP;Daniels,L;Orme-Johnson,WH;Walsh,C
产甲烷细菌Methanobacterium thermoautotrophicum菌株ΔH中的两种氢化酶已被纯化，它们含有紧密结合的镍以及预期的铁/硫原子，铁/硫原子的固定比例为15-20个铁原子/镍。一种氢化酶还原8-羟基-5-脱氮黄素辅酶因子420（F420），而另一种已被纯化为甲基紫还原氢化酶。这两种酶都具有顺磁性镍的EPR信号，如在含61镍氢化酶中的超精细偶联所示。与模型化合物的比较表明这些有氧纯化的酶的非活性形式中的镍（III）氧化态。镍（III）信号的损失伴随着还原活化，但不与高比活度的恢复动力学相关。在气相中用氩气取代H2时，在还原的、活性的F420还原酶上，出现了几个EPR信号，包括可能是酶结合的FADH半醌的信号atg= 2.004，反映顺磁性镍（III）的新形式的两个信号atg= 2.140和2.196，以及可能是铁信号的信号atg= 2.036。第二顺磁性镍形式的F420还原氢化酶本身是活性的，或者与活性酶处于容易的平衡。在g = 2.036处的信号的大小可能与酶的活化程度相关。与Clostridium pasteurianum[Erbes，D. L.，伯里斯河H. & Orme-Johnson，W. H.等人（1975）Proc. Acad. Sci. USA 72，4795-4799]，似乎仅使用铁/硫辅基并与单电子转移剂反应，这种产甲烷菌氢化酶似乎利用铁、镍和黄素氧化还原位点并还原专性单电子（紫精）和双电子（脱氮黄霉素）氧化剂。
Two hydrogenases from the methanogenic bacteriumMethanobacterium thermoautotrophicumstrain ΔH have been purified and contain tightly bound nickel as well as the anticipated iron/sulfur atoms with a fixed ratio of 15-20 iron atoms per nickel. One hydrogenase reduces the 8-hydroxy-5-deazaflavin coenzyme factor 420 (F420), whereas the other has been purified as a methyl viologen-reducing hydrogenase. Both enzymes possess an EPR signal attributed to paramagnetic nickel as demonstrated by hyperfine coupling in61Ni-containing hydrogenases. Comparison to model compounds suggests a nickel(III) oxidation state in the inactive forms of these aerobically purified enzymes. Loss of the nickel(III) signal accompanies reductive activation but is not kinetically correlated with regain of high specific activity. On replacement of H2by argon in the gas phase over reduced, active, F420-reducing enzyme, several EPR signals appear, including a signal atg= 2.004 that is probably enzyme-bound FADH semiquinone, two signals atg= 2.140 and 2.196 that reflect a new form of paramagnetic nickel(III), and also a signal atg= 2.036 that may be an iron signal. The F420-reducing hydrogenase in the second paramagnetic nickel form is either itself active or in facile equilibrium with active enzyme. The size of the signal atg= 2.036 may correlate with the degree of activation of the enzyme. In contrast to the hydrogenase ofClostridium pasteurianum[Erbes, D. L., Burris, R. H. & Orme-Johnson, W. H. (1975)Proc. Natl. Acad. Sci. USA72, 4795-4799], which appears to use only iron/sulfur prosthetic groups and which reacts with one-electron-transfer agents, this methanogen hydrogenase seems to utilize iron, nickel, and flavin redox sites and to reduce obligate one-electron (viologen) and two-electron (deazaflavin) oxidants.