菜心中高等电点高活性的乙醇酸氧化酶同工酶的纯化和特性  

Purification and Characterization of Glycolate Oxidase Isozyme with High Isoelectric Point and High Activity from Green Leaves of Brassica parachinensis Bailey

作　　者：尹汉萍[1,2] 徐杰[1,2] 曾秋莲[1] 王再花[1] 叶庆生[1] 董宇亮[1] 黄美意[1] 韩雪[1] 苏燕琼[1] 庄莹莹[1] 

机构地区：[1]华南师范大学生命科学学院,广东省植物发育生物工程重点实验室 [2]中国农业大学植物生理学与生物化学国家重点开放实验室,北京100094

出　　处：《中国生物化学与分子生物学报》2004年第5期690-695,共6页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金 (No .3 980 0 0 0 9);中国农业大学植物生理学与生物化学国家重点开放实验室基金 (G19980 10 10 0 8和G19990 1170 0 4);广东省自然科学基金 (No.0 2 110 5 )资助~~

摘　　要：Glycolate oxidase (GO) isozyme with high specific activity (75.0～279.0 U/mg) is purified quickly on DEAE- Cellulose column from Brassica parachinensis Bailey. Its pI is greater than 10.0 assayed by acetate cellulose membrane electrophoresis for 1 hour. In view of about ten kinds of pI varied from 4.5 to 10.0 are observed when the same GO isozyme is assayed in IEF for 14 hours, it is obvious that its pI decreases in IEF. Its pI also decreases when this GO isozyme is assayed in PAGE for 14 hours. Based on the results in SDS-PAGE, CGE-SDS, and IEF, it is most likely that this GO isozyme comprises two noncovalently associated 66 kD basic subunit and 40 kD acidic subunit, the phenomenon of pI change is related to subunit dissociation. The basic/acidic amino acid residues ratios in GO isozyme and its 40 kD acidic subunit are detected to be 0.66 and 0.54, respectively, a value much lower than that (0.96) in 40 kD peptide encoded by GO cDNA reported previously, indicating neither M r nor charge characteristic of this 40 kD peptide is similar to that of GO isozyme subunits, two subunits of GO isozyme may be the modified products of the same GO gene after post-translation.Glycolate oxidase (GO) isozyme with high specific activity (75.0～279.0 U/mg) is purified quickly on DEAE- Cellulose column from Brassica parachinensis Bailey. Its pI is greater than 10.0 assayed by acetate cellulose membrane electrophoresis for 1 hour. In view of about ten kinds of pI varied from 4.5 to 10.0 are observed when the same GO isozyme is assayed in IEF for 14 hours, it is obvious that its pI decreases in IEF. Its pI also decreases when this GO isozyme is assayed in PAGE for 14 hours. Based on the results in SDS-PAGE, CGE-SDS, and IEF, it is most likely that this GO isozyme comprises two noncovalently associated 66 kD basic subunit and 40 kD acidic subunit, the phenomenon of pI change is related to subunit dissociation. The basic/acidic amino acid residues ratios in GO isozyme and its 40 kD acidic subunit are detected to be 0.66 and 0.54, respectively, a value much lower than that (0.96) in 40 kD peptide encoded by GO cDNA reported previously, indicating neither M r nor charge characteristic of this 40 kD peptide is similar to that of GO isozyme subunits, two subunits of GO isozyme may be the modified products of the same GO gene after post-translation.

关 键 词：乙醇酸氧化酶 酶的纯化 IEF 等电点 CGE 同工酶 DNA 菜心 SDS-PAGE 特性 

分 类 号：Q949.782[生物学—植物学] S688.4[农业科学—观赏园艺]