CLEAVAGE OF TRANSFER-RNA WITH IMIDAZOLE AND SPERMINE IMIDAZOLE CONSTRUCTS - A NEW APPROACH FOR PROBING RNA STRUCTURE

被引：93

作者：

VLASSOV, VV

ZUBER, G

FELDEN, B

BEHR, JP

GIEGE, R

机构：

[1] INST BIOL MOLEC & CELLULAIRE,CNRS,UPR 9002,F-67084 STRASBOURG,FRANCE

[2] RUSSIAN ACAD SCI,INST BIOORGAN CHEM,NOVOSIBIRSK 630090,RUSSIA

[3] NOVOSIBIRSK STATE UNIV,NOVOSIBIRSK 630090,RUSSIA

[4] UNIV LOUIS PASTEUR STRASBOURG 1,FAC PHARM,CHIM GENET LAB,CNRS,URA 1386,F-67401 ILLKIRCH GRAFFENS,FRANCE

来源：

NUCLEIC ACIDS RESEARCH | 1995年 / 23卷 / 16期

关键词：

D O I：

10.1093/nar/23.16.3161

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：

Hydrolysis of RNA in imidazole buffer and by spermine-imidazole conjugates has been investigated. The RNA models were yeast tRNA(Asp) and a transcript derived from the 3'-terminal sequence of tobacco mosaic virus RNA representing a minihelix capable of being enzymatically aminoacylated with histidine. Imidazole buffer and spermine-imidazole conjugates in the presence of free imidazole cleave phosphodiester bonds in the folded RNAs in a specific fashion. Imidazole buffer induces cleavages preferentially in single-stranded regions because nucleotides in these regions have more conformational freedom and can assume more easily the geometry needed for formation of the hydrolysis intermediate state. Spermine-imidazole constructs supplemented with free imidazole cleave tRNA(Asp) within single-stranded regions after pyrimidine residues with a marked preference for pyrimidine-A sequences. Hydrolysis patterns suggest a cleavage mechanism involving an attack by the imidazole residue of the electrostatically bound spermine-imidazole and by free imidazole at the most accessible single-stranded regions of the RNA. Cleavages in a viral RNA fragment recapitulating a tRNA-like domain were found in agreement with the model of this molecule that accounts for its functional properties, thus illustrating the potential of the imidazole-derived reagents as structural probes for solution mapping of RNAs. The cleavage reactions are simple to perform, provide information reflecting the state of the ribose-phosphate backbone of RNA and can be used for mapping single-and double-stranded regions in RNAs.

引用

页码：3161 / 3167

页数：7

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