A di-heme cytochrome c peroxidase from Nitrosomonas europaea catalytically active in both the oxidized and half-reduced states.
A di-heme cytochrome c peroxidase from Nitrosomonas europaea catalytically active in both the oxidized and half-reduced states.
复制标题
一种来自欧洲亚硝化单胞菌的二血红素细胞色素 c 过氧化物酶，在氧化和半还原状态下均具有催化活性。
DOI：
--
复制
发表时间：
1994
影响因子：
4.8
通讯作者：
A. Hooper
中科院分区：
文献类型：
--
作者：
D. Arciero;A. Hooper
从趋化自养细菌europaea亚硝化单胞菌中分离到一种含细胞色素（细胞色素c553过氧化物酶）的二c-血红素。对N端和两种含血红素肽的序列分析显示，与铜绿假单胞菌中二血红素过氧化物酶的序列总体上有40%的同源性（Rönnberg， M., Kalkkinen， N., and Ellfolk， N. (1989) FEBS Lett. 250, 175-178）。在室温和pH 7.0条件下，一个血红素为低自旋Em7 = +450 mV，另一个血红素为高自旋Em7 = -260 mV。低温下EPR谱显示出高自旋和低自旋信号的混合。当细胞色素c553过氧化物酶完全氧化（FeLS3+:FeHS3+）或半还原（FeLS2+:FeHS3+）时，阴离子配体（CN-, N3-, F-, CNO-）结合从而扰乱高自旋血红素。在完全氧化的酶中，高电位、低自旋血红素的EPR信号不受配体存在的干扰。此外，对于过氧化物酶的任一氧化态，每种配体都会产生相似的特征EPR信号。细胞色素c553过氧化物酶的完全氧化和半还原氧化状态都具有催化活性，这可以通过酶在H2O2存在下氧化马心脏细胞色素c的能力以及过氧化氢加入过氧化物酶后的光学变化来证明。在化学计量量的H2O2存在下，半还原酶被迅速氧化，完全氧化的酶在光谱Soret区域的吸光度显著下降，在600-650 nm附近有较小的增加。后一种光学变化类似于在卟啉阳离子自由基形成过程中观察到的变化。这表明这种二血红素过氧化物酶可能形成类似于辣根过氧化物酶形成的化合物I中间体。
A di-c-heme containing cytochrome (cytochrome c553 peroxidase) has been isolated from the chemoautotrophic bacterium Nitrosomonas europaea. Sequence analysis of the N terminus and the two heme-containing peptides generated by digestion of the enzyme with trypsin show 40% homology overall to sequences reported for the di-heme peroxidase from Pseudomonas aeruginosa (Rönnberg, M., Kalkkinen, N., and Ellfolk, N. (1989) FEBS Lett. 250, 175-178). At room temperature and pH 7.0, one heme is low spin with Em7 = +450 mV and the other is high spin with Em7 = -260 mV. EPR spectra show a mixture of high spin and low spin signals at cryogenic temperatures. Anionic ligands (CN-, N3-, F-, CNO-) bind so as to perturb the high spin heme when cytochrome c553 peroxidase is either fully oxidized (FeLS3+:FeHS3+) or half-reduced (FeLS2+:FeHS3+). The EPR signal of the high potential, low spin heme in fully oxidized enzyme is unperturbed by the presence of the ligands. Furthermore, each ligand results in similar characteristic EPR signals for either oxidation state of the peroxidase. Both the fully oxidized and half-reduced oxidation states of cytochrome c553 peroxidase are catalytically active as evidenced by the enzyme's ability to oxidize horse heart cytochrome c in the presence of H2O2, as well as by optical changes associated with the addition of H2O2 to the peroxidase. In the presence of stoichiometric amounts of H2O2, the half-reduced enzyme is rapidly oxidized and the fully oxidized enzyme shows a significant decrease in absorbance in the Soret region of the optical spectrum coupled with a lesser increase near 600-650 nm. These latter optical changes are similar to what is observed in the formation of a porphyrin cation radical. This suggests that this di-heme peroxidase may form a compound I intermediate analogous to that formed by horseradish peroxidase.