Primus J L, Boersma M G, Mandon D, Boeren S, Veeger C, Weiss R, Rietjens I M
Laboratory of Biochemistry, Agricultural University, Wageningen, The Netherlands.
J Biol Inorg Chem. 1999 Jun;4(3):274-83. doi: 10.1007/s007750050313.
This study describes the catalytic properties of manganese microperoxidase 8 [Mn(III)MP8] compared to iron microperoxidase 8 [Fe(III)MP8]. The mini-enzymes were tested for pH-dependent activity and operational stability in peroxidase-type conversions, using 2-methoxyphenol and 3,3'-dimethoxybenzidine, and in a cytochrome P450-like oxygen transfer reaction converting aniline to para-aminophenol. For the peroxidase type of conversions the Fe to Mn replacement resulted in a less than 10-fold decrease in the activity at optimal pH, whereas the aniline para-hydroxylation is reduced at least 30-fold. In addition it was observed that the peroxidase type of conversions are all fully blocked by ascorbate and that aniline para-hydroxylation by Fe(III)MP8 is increased by ascorbate whereas aniline para-hydroxylation by Mn(III)MP8 is inhibited by ascorbate. Altogether these results indicate that different types of reactive metal oxygen intermediates are involved in the various conversions. Compound I/II, scavenged by ascorbate, may be the reactive species responsible for the peroxidase reactions, the polymerization of aniline and (part of) the oxygen transfer to aniline in the absence of ascorbate. The para-hydroxylation of aniline by Fe(III)MP8, in the presence of ascorbate, must be mediated by another reactive iron-oxo species which could be the electrophilic metal(III) hydroperoxide anion of microperoxidase 8 [M(III)OOH MP8]. The lower oxidative potential of Mn, compared to Fe, may affect the reactivity of both compound I/II and the metal(III) hydroperoxide anion intermediate, explaining the differential effect of the Fe to Mn substitution on the pH-dependent behavior, the rate of catalysis and the operational stability of MP8.
本研究描述了锰微过氧化物酶8 [Mn(III)MP8]与铁微过氧化物酶8 [Fe(III)MP8]相比的催化特性。使用2-甲氧基苯酚和3,3'-二甲氧基联苯胺，对这些微型酶在过氧化物酶型转化中的pH依赖性活性和操作稳定性进行了测试，并在将苯胺转化为对氨基苯酚的细胞色素P450样氧转移反应中进行了测试。对于过氧化物酶型转化，Fe被Mn取代导致在最佳pH下活性降低不到10倍，而苯胺对羟基化至少降低30倍。此外，观察到过氧化物酶型转化均被抗坏血酸完全阻断，并且Fe(III)MP8催化的苯胺对羟基化被抗坏血酸增强，而Mn(III)MP8催化的苯胺对羟基化被抗坏血酸抑制。总之，这些结果表明不同类型的活性金属氧中间体参与了各种转化。被抗坏血酸清除的化合物I/II可能是负责过氧化物酶反应、苯胺聚合以及在没有抗坏血酸时向苯胺的（部分）氧转移的活性物种。在抗坏血酸存在下，Fe(III)MP8催化的苯胺对羟基化必须由另一种活性铁氧物种介导，该物种可能是微过氧化物酶8的亲电金属(III)氢过氧化物阴离子[M(III)OOH MP8]。与Fe相比，Mn的较低氧化电位可能会影响化合物I/II和金属(III)氢过氧化物阴离子中间体的反应性，这解释了Fe被Mn取代对MP8的pH依赖性行为、催化速率和操作稳定性的不同影响。
