A highly efficient procedure for purifying the ribosome-inactivating proteins alpha- and beta-momorcharins from Momordica charantia seeds, N-terminal sequence comparison and establishment of their N-glycosidase activity

被引:37
作者
Fong, WP [1 ]
Poon, YT [1 ]
Wong, TM [1 ]
Mock, JWY [1 ]
Ng, TB [1 ]
Wong, RNS [1 ]
Yao, QZ [1 ]
Yeung, HW [1 ]
机构
[1] HONG KONG BAPTIST UNIV, DEPT BIOL, KOWLOON, HONG KONG
关键词
purification; Momordica charantia; ribosome-inactivating proteins; RNA; N-glycosidase;
D O I
10.1016/0024-3205(96)00388-8
中图分类号
R-3 [医学研究方法]; R3 [基础医学];
学科分类号
1001 ;
摘要
A new purification scheme, involving two successive ion exchange chromatographic steps on DEAF-cellulose and Mono-S FPLC, was developed for the isolation of the ribosome-inactivating proteins, alpha- and beta-momorcharins, from the Chinese herb Kuquazi (seeds of Momordica charantia). This simple and rapid procedure yielded 3.1 and 1.7 mg of alpha- and beta-momorcharins, respectively, from 2.5 g of decorticated seeds in only two days. The N-terminal amino acid sequence of beta-momorcharin was found to be DVNFDLSTATAKTYTKFIED. It differed from that of alpha-momorcharin (DVSFRLSGADPRSYGMFIKD) in 10 out of the 20 positions investigated. Like other ribosome-inactivating proteins, the purified momorcharins showed specific N-glycosidase activity at nanomolar concentrations, when rRNA from rabbit reticulocyte lysate was used as substrate. The N-glycosidase activity of both momorcharins was optimal at pH7, not inhibited by K+ and not appreciably affected by NH4+. The activity of alpha-momorcharin was not drastically altered by Mn2+ but (1-10mM) Mn2+ inhibited the activity of beta-momorcharin by about 40%.
引用
收藏
页码:901 / 909
页数:9
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