The coprophilous mushroom Coprinus radians secretes a haloperoxidase that catalyzes aromatic peroxygenation
The coprophilous mushroom Coprinus radians secretes a haloperoxidase that catalyzes aromatic peroxygenation
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DOI：
10.1128/aem.00026-07
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发表时间：
2007-09-01
影响因子：
4.4
通讯作者：
Hofrichter, Martin
中科院分区：
文献类型：
--
作者：
Anh, Dau Hung;Ullrich, Rene;Hofrichter, Martin
采用选择性琼脂平板法和豆粕复合培养基法对26株鬼伞属粪粪腐菌进行过氧化物酶活性筛选。本文报道了两个种：Coprinus radians和C.发现轮枝藻产生过氧化物酶，其在pH 7下将芳基醇氧化成相应的醛（血红素-硫醇盐卤代过氧化物酶的典型反应）。对鸡腿菇过氧化物酶进行了纯化和表征。三个馏分的酶，磷酸肌酸I，磷酸肌酸II，磷酸肌酸III，分子量为43至45 kDa，以及等电点之间的3.8和4.2，被确定后，通过阴离子交换和尺寸排阻色谱纯化。主要组分（CrP 11）氧化芳基醇的最适pH为7左右，就酶活性和稳定性而言，0.7 mM的H2 O2浓度是最合适的。ABTS [2，2 '-连氮双（3-乙基苯并噻唑啉磺酸）]、2，6-二甲氧基苯酚、苯甲醇、藜芦醇和H2 O2的表观K-m值分别为49、342、635、88和1，201 μ M。N端与茶树菇过氧化物酶（Agrocybe aegerita peroxidase，AaP？）和氯过氧化物酶。CrP II的紫外-可见光谱与静息态细胞色素P450酶的紫外-可见光谱高度相似，Soret带位于422 nm，另外的最大值位于359、542和571 nm。还原的一氧化碳复合物在446 nm处显示出最大吸收，这是血红素硫醇盐蛋白的特征。CrP溴化苯酚为2-和4-溴苯酚，选择性羟基化萘为I-萘酚。因此，在AaP之后，CrP是迄今为止描述的第二种细胞外卤代过氧化物酶-过氧化物酶。胞外羟基化芳香族化合物的能力似乎是关键的催化性能的磷酸肌酸，并可能是一般意义的生物转化的难利用的芳香族物质，如木质素，腐殖质，和有机污染物在土壤凋落物和粪便环境。此外，芳香族过氧化是生物技术研究的一个有前途的目标。
Coprophilous and litter-decomposing species (26 strains) of the genus Coprinus were screened for peroxidase activities by using selective agar plate tests and complex media based on soybean meal. Two species, Coprinus radians and C. verticillatus, were found to produce peroxidases, which oxidized aryl alcohols to the corresponding aldehydes at pH 7 (a reaction that is typical for heme-thiolate haloperoxidases). The peroxidase of Coprinus radians was purified to homogeneity and characterized. Three fractions of the enzyme, CrP I, CrP II, and CrP III, with molecular masses of 43 to 45 kDa as well as isoelectric points between 3.8 and 4.2, were identified after purification by anion-exchange and size exclusion chromatography. The optimum pH of the major fraction (CrP 11) for the oxidation of aryl alcohols was around 7, and an H2O2 concentration of 0.7 mM was most suitable regarding enzyme activity and stability. The apparent K-m values for ABTS [2,2'-azinobis(3-ethylbenzthiazolinesulfonic acid)], 2,6-dimethoxyphenol, benzyl alcohol, veratryl alcohol, and H2O2 were 49, 342, 635, 88, and 1,201 mu M, respectively. The N terminus of CrP II showed 29% and 19% sequence identity to Agrocybe aegerita peroxidase (AaP?) and chloroperoxidase, respectively. The UV-visible spectrum of CrP II was highly similar to that of resting-state cytochrome P450 enzymes, with the Soret band at 422 urn and additional maxima at 359, 542, and 571 nm. The reduced carbon monoxide complex showed an absorption maximum at 446 nm, which is characteristic of heme-thiolate proteins. CrP brominated phenol to 2- and 4-bromophenols and selectively hydroxylated naphthalene to I-naphthol. Hence, after AaP, CrP is the second extracellular haloperoxidase-peroxygenase described so far. The ability to extracellularly hydroxylate aromatic compounds seems to be the key catalytic property of CrP and may be of general significance for the biotransformation of poorly available aromatic substances, such as lignin, humus, and organopollutants in soil litter and dung environments. Furthermore, aromatic peroxygenation is a promising target of biotechnological studies.