在甲基营养型酵母毕赤酵母中高水平生产菠菜乙醇酸氧化酶：构建生物催化剂

High-level production of spinach glycolate oxidase in the methylotrophic yeast Pichia pastoris: engineering a biocatalyst.

作者信息

Payne M S, Petrillo K L, Gavagan J E, Wagner L W, DiCosimo R, Anton D L

机构信息

DuPont Central Research and Development, Glasgow Site, Newark, DE 19714-6101, USA.

出版信息

Gene. 1995 Dec 29;167(1-2):215-9. doi: 10.1016/0378-1119(95)00661-3.

DOI:10.1016/0378-1119(95)00661-3

PMID:8566780

Abstract

Glycolate oxidase (GO) is a flavo-enzyme that catalyzes the oxidation of glycolate, and is useful for the biocatalytic production of glyoxylate. We have produced high levels of spinach GO in the methylotrophic yeast Pichia pastoris (Pp), by chromosomal integration of multiple copies of an expression cassette containing the GO coding sequence under control of the methanol-inducible alcohol oxidase I promoter. Under fermentation conditions, greater than 250 units of GO per gram of cells (wet weight) was obtained, corresponding to roughly 20-30% of soluble cell protein. This recombinant Pp strain was used as a whole-cell biocatalyst for conversion of glycolic acid to glyoxylic acid.

摘要

乙醇酸氧化酶（GO）是一种黄素酶，可催化乙醇酸的氧化，对生物催化生产乙醛酸很有用。我们通过在甲醇诱导型醇氧化酶I启动子控制下，将含有GO编码序列的表达盒的多个拷贝进行染色体整合，在甲基营养型酵母毕赤酵母（Pp）中高水平表达了菠菜GO。在发酵条件下，每克细胞（湿重）可获得超过250个单位的GO，约占可溶性细胞蛋白的20 - 30%。该重组Pp菌株用作全细胞生物催化剂，将乙醇酸转化为乙醛酸。