聚乙二醇衍生物修饰的胆固醇氧化酶在苯中可溶且具有活性。

Polyethylene glycol derivative-modified cholesterol oxidase soluble and active in benzene.

作者信息

Yoshimoto T, Ritani A, Ohwada K, Takahashi K, Kodera Y, Matsushima A, Saito Y, Inada Y

机构信息

Laboratory of Biological Chemistry, Tokyo Institute of Technology, Japan.

出版信息

Biochem Biophys Res Commun. 1987 Oct 29;148(2):876-82. doi: 10.1016/0006-291x(87)90957-0.

DOI:10.1016/0006-291x(87)90957-0

PMID:3479986

Abstract

Cholesterol oxidase from Nocardia sp. was modified with a synthetic copolymer of polyoxyethylene allylmethyldiether (PEG) and maleic acid anhydride (MA anhydride), poly(PEG-MA anhydride). The modified cholesterol oxidase, in which 64% of the amino groups in the protein molecule were coupled to poly(PEG-MA), was soluble in organic solvents and catalyzed the oxidation reaction of cholesterol in benzene to form 4-cholesten-3-one with the enzymic activity of 0.6 mumol/min/mg protein. Using the modified cholesterol oxidase together with polyethylene glycol-modified peroxidase, coupled reactions shown below took place in Cholesterol + O2----4-Cholesten-3-one + H2O2 H2O2 + o-Phenylenediamine----H2O + Oxidized o-Phenylenediamine transparent benzene solution, not in an emulsified system. The oxidation of cholesterol was directly determined in benzene by measuring the absorbance of oxidized o-phenylenediamine at 490 nm.

摘要

来自诺卡氏菌属的胆固醇氧化酶用聚氧乙烯烯丙基甲基二醚（PEG）和马来酸酐（MA酐）的合成共聚物聚（PEG-MA酐）进行修饰。修饰后的胆固醇氧化酶中，蛋白质分子中64%的氨基与聚（PEG-MA）偶联，可溶于有机溶剂，并催化胆固醇在苯中的氧化反应生成4-胆甾烯-3-酮，酶活性为0.6 μmol/分钟/毫克蛋白质。将修饰后的胆固醇氧化酶与聚乙二醇修饰的过氧化物酶一起使用时，在胆固醇 + O₂→4-胆甾烯-3-酮 + H₂O₂ 、H₂O₂ + 邻苯二胺→H₂O + 氧化型邻苯二胺的反应中，是在透明的苯溶液中而非乳化体系中发生如下偶联反应。通过测量氧化型邻苯二胺在490 nm处的吸光度，可直接在苯中测定胆固醇的氧化情况。