Intrinsic ribonuclease activities in ribonuclease and ribosome-inactivating proteins from the seeds of bitter gourd

被引:40
作者
Fong, WP [1 ]
Mock, WY [1 ]
Ng, TB [1 ]
机构
[1] Chinese Univ Hong Kong, Dept Biochem, Shatin, Hong Kong, Peoples R China
关键词
momorcharin; Momordica charantia seeds; purification; ribonuclease; ribosome-inactivating proteins;
D O I
10.1016/S1357-2725(99)00149-1
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Alpha- and beta-momorcharins are ribosome-inactivating proteins present in the seeds of the bitter gourd (Momordica charantia). Both of them possess ribonuclease activity which may account for some of their biological properties. However, the activity is weak and hence it is important to confirm that the ribonuclease activity observed is not due to any contamination. To this end, the ribonuclease from the seeds of M. charantia (RNase-MC) was purified and compared with the ribonuclease activity of the momorcharins. Purification was achieved by ion-exchange chromatographies on DEAE-cellulose, SP-Sepharose and Mono-S. RNase-MC had a molecular mass of 22 kDa. It acted on tRNA to release acid-soluble UV-absorbing species with a pH optimum around 6.0 6.5. When polyhomoribonucleotides were used as substrates, it was found that RNase-MC acted preferentially on polyU but exerted much weaker activity on polyC, polyG and polyA. Chromatographic analysis of the reaction product indicated that mono- and oligo-ribonucleotides, but not free base, were generated from polyU, suggesting that the enzymatic action involved ribonucleolytic cleavage. RNase-MC exhibited a much more potent (at least 1000-fold higher) ribonuclease activity than alpha- and beta-momorcharins. RNase-MC, alpha-momorcharin and beta-momorcharin were separable on Mono-S, indicating that the ribonuclease activities present in the three proteins were distinct entities. (C) 2000 Elsevier Science Ltd. All rights reserved.
引用
收藏
页码:571 / 577
页数:7
相关论文
共 22 条
[1]  
BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3
[2]   The deoxyribonuclease activity attributed to ribosome-inactivating proteins is due to contamination [J].
Day, PJ ;
Lord, JM ;
Roberts, LM .
EUROPEAN JOURNAL OF BIOCHEMISTRY, 1998, 258 (02) :540-545
[3]   CRYSTALLIZATION AND PRELIMINARY-X-RAY DIFFRACTION ANALYSIS OF A PLANT RIBONUCLEASE FROM THE SEEDS OF THE BITTER GOURD MOMORDICA-CHARANTIA [J].
DE, A ;
FUNATSU, G .
JOURNAL OF MOLECULAR BIOLOGY, 1992, 228 (04) :1271-1273
[4]   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 [J].
Fong, WP ;
Poon, YT ;
Wong, TM ;
Mock, JWY ;
Ng, TB ;
Wong, RNS ;
Yao, QZ ;
Yeung, HW .
LIFE SCIENCES, 1996, 59 (11) :901-909
[5]   DEOXYRIBONUCLEOLYTIC ACTIVITY OF ALPHA-MOMORCHARIN AND BETA-MOMORCHARIN [J].
GO, TTM ;
YEUNG, HW ;
FONG, WP .
LIFE SCIENCES, 1992, 51 (17) :1347-1353
[6]  
IDE H, 1991, FEBS LETT, V289, P126, DOI 10.1016/0014-5793(91)80924-R
[7]   THE COMPLETE AMINO-ACID-SEQUENCE OF RIBONUCLEASE FROM THE SEEDS OF BITTER GOURD (MOMORDICA-CHARANTIA) [J].
IDE, H ;
KIMURA, M ;
ARAI, M ;
FUNATSU, G .
FEBS LETTERS, 1991, 284 (02) :161-164
[8]   AMINO-ACID-SEQUENCE OF AN EXTRACELLULAR, PHOSPHATE-STARVATION-INDUCED RIBONUCLEASE FROM CULTURED TOMATO (LYCOPERSICON-ESCULENTUM) CELLS [J].
JOST, W ;
BAK, H ;
GLUND, K ;
TERPSTRA, P ;
BEINTEMA, JJ .
EUROPEAN JOURNAL OF BIOCHEMISTRY, 1991, 198 (01) :1-6
[9]   MATURATION OF HEAD OF BACTERIOPHAGE-T4 .1. DNA PACKAGING EVENTS [J].
LAEMMLI, UK ;
FAVRE, M .
JOURNAL OF MOLECULAR BIOLOGY, 1973, 80 (04) :575-599
[10]   Purification and characterization of novel ribosome inactivating proteins, alpha- and beta-pisavins, from seeds of the garden pea Pisum sativum [J].
Lam, SSL ;
Wang, HX ;
Ng, TB .
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 1998, 253 (01) :135-142