利用毕赤酵母表达系统纯化重组真核单胺氧化酶A和单胺氧化酶B

Purification of Recombinant Eukaryotic MAO A and MAO B Utilizing the Pichia pastoris Expression System.

作者信息

Edmondson Dale E

机构信息

Department of Biochemistry, Emory University, Atlanta, GA, USA.

出版信息

Methods Mol Biol. 2023;2558:11-22. doi: 10.1007/978-1-0716-2643-6_2.

DOI:10.1007/978-1-0716-2643-6_2

PMID:36169852

Abstract

Procedures are described for the heterologous expression and purification of the mitochondrial-bound enzymes human and rat monoamine oxidases A and B and zebrafish MAO in the yeast Pichia pastoris. Enzyme expression is under control of a methanol oxidase promoter and similar procedures have been developed for the preparation of membrane particles and detergent solubilization of the functional enzymes. Similarities and differences are described in the procedures for purification of the respective enzymes using standard column chromatographic techniques to provide enzyme yields in the range of 100-300 mg from 1 L of cell culture.

摘要

本文描述了在毕赤酵母中异源表达和纯化人源、大鼠源单胺氧化酶A和B以及斑马鱼单胺氧化酶（MAO）这几种线粒体结合酶的方法。酶的表达受甲醇氧化酶启动子的控制，并且已经开发了类似的方法用于制备膜颗粒以及功能性酶的去污剂增溶。文中还描述了使用标准柱色谱技术纯化各酶的方法中的异同点，以从1升细胞培养物中获得产量在100 - 300毫克范围内的酶。