烟曲霉中烟曲霉氯过氧化物酶的表达及重组酶的特性研究
Expression of the Caldariomyces fumago chloroperoxidase in Aspergillus niger and characterization of the recombinant enzyme.
作者信息
Conesa A, van De Velde F, van Rantwijk F, Sheldon R A, van Den Hondel C A, Punt P J
机构信息
Department of Applied Microbiology and Gene Technology, TNO Nutrition and Food Research Institute, Utrechtseweg 48, 3704 HE Zeist, The Netherlands.
出版信息
J Biol Chem. 2001 May 25;276(21):17635-40. doi: 10.1074/jbc.M010571200. Epub 2001 Feb 22.
The Caldariomyces fumago chloroperoxidase was successfully expressed in Aspergillus niger. The recombinant enzyme was produced in the culture medium as an active protein and could be purified by a three-step purification procedure. The catalytic behavior of recombinant chloroperoxidase (rCPO) was studied and compared with that of native CPO. The specific chlorination activity (47 units/nmol) of rCPO and its pH optimum (pH 2.75) were very similar to those of native CPO. rCPO catalyzes the oxidation of various substrates in comparable yields and selectivities to native CPO. Indole was oxidized to 2-oxindole with 99% selectivity and thioanisole to the corresponding R-sulfoxide (enantiomeric excess >98%). Incorporation of (18)O from labeled H(2)18O(2) into the oxidized products was 100% in both cases.
烟曲霉氯过氧化物酶在黑曲霉中成功表达。重组酶在培养基中作为活性蛋白产生，可通过三步纯化程序进行纯化。对重组氯过氧化物酶（rCPO）的催化行为进行了研究，并与天然CPO进行了比较。rCPO的比氯化活性（47单位/纳摩尔）及其最适pH值（pH 2.75）与天然CPO非常相似。rCPO催化各种底物氧化的产率和选择性与天然CPO相当。吲哚以99%的选择性氧化为2-氧代吲哚，苯甲硫醚氧化为相应的R-亚砜（对映体过量>98%）。在这两种情况下，标记的H₂¹⁸O₂中的¹⁸O掺入氧化产物的比例均为100%。
Zotero 插件