[Complementary addressed alkylation of Escherichia coli 16S rRNA with 2',3'-O-[4-N-methyl-N-(2-chloroethyl)aminobenzylidene]m derivatives of oligodeoxyribonucleotides. IV. Determination of binding sites of 16S rRNA with benzylidene derivatives of d(pACCTTGTT)rA, d(pTTACGACT)rU, d(TTTGCTCCCC)rA].
By site-directed alkylation of 16S rRNA with benzylidene derivatives of d(pACCTTGTT)rA (II), d(pTTACGACT)rU (III), d(pTTTGCTCCCC)rA (IV) (reagents (II)--(IV] followed by the RNase H treatment a number of 16S rRNA fragments have been obtained. Hybridisation of these fragments with restriction fragments of plasmid pKK 3535, containing operon rrnB of E. coli rRNAs, led to the identification of all reagents' binding sites in 16S rRNA. Good correlation is found between estimated stability of non-perfect 16S rRNA.oligodeoxyribonucleotide duplexes and the level of modification of this site with alkylating derivative of the same oligodeoxyribonucleotide. With high concentration of the reagents (II)--(IV) ((2-5) x 10(-5) M) the site-directed alkylation proceeds not only at the desired site but also at other sites corresponding to non-perfect duplexes between 16S rRNA and the reagents. It should be noted that the modification mainly occurs in the non-perfect duplexes, carrying mismatched bases at the termini. Influence of the secondary structure of 16S rRNA on the site-directed modification is discussed.
Zenkova
M A
MA
Karpova
G G
GG
Levina
A S
AS
Mamaev
S V
SV
Solov'ev
V V
VV
D000478:Alkylation; D001483:Base Sequence; D001597:Benzylidene Compounds; D001665:Binding Sites; D004926:Escherichia coli / Q000378:metabolism*; D006868:Hydrolysis; D007202:Indicators and Reagents; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000378:metabolism*; D012336:RNA, Ribosomal, 16S / Q000378:metabolism*
D001597:Benzylidene Compounds; D007202:Indicators and Reagents; D009838:Oligodeoxyribonucleotides; D012336:RNA, Ribosomal, 16S
Komplementarno adresovannoe alkilirovanie 16S rRNK Escherichia coli 2',3'-O-[4-methyl-N-(2-khlorétil)aminobenzilidenovymi] proizvodnymi oligodezoksiribonukleotidov. IV. Opredelenie uchastkov sviazyvaniia 16S rRNK s benzilidenovymi proizvodnymi d(pACCTTGTT)rA, d(pTTACGACT)rU, d(TTTGCTCCCC)rA.