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Nucleic Acids Research, 1989, Vol. 17, No. 19 7833-7842
© 1989

CHEMISTRY

Deoxynucleotide-containing oligoribonuccleotide duplexes: stability and susceptibility to RNase V1 and RNase H

Jacqueline R. Wyatt and G.Terrance Walker*

Department of Chemistry and Laboratory of Chemical Biodynamics, University of California Berkeley, CA 94720, USA

*Present address: Becton-Dickinson Research Center, PO Box 12016, Research Triangle Park, NC 27709, USA

Received May 4, 1989. Revised September 1, 1989. Accepted September 1, 1989.

Oligoribonucleotide duplexes containing one to four 2'-deoxynucleotide residues were used as substrates for ribonuclease V1 and RNase H. Either deoxyadenosine and/or deoxythymidine were incorporated into the duplex, 5'GGCCGGAUCCGCGC3'·5'GCGCGGAUCCGGCC3' by substitution of the appropriate deoxynucleoside triphosphate into a transcription reaction with T7 RNA polymerase. The melting temperature, Tm, of the duplex (1.8 µM in strands in 50 mM NaCl) containing only ribonucleotides was 79.9°C. Substitution of deoxyadenosine in both strands of the duplex lowered the Tm by 2.4°C. Substitution of deoxythymidine had no measurable effect on the Tm. Comparison of RNase V1 digestion patterns of fully ribonucleotide and deoxy-substituted duplexes suggest that any distortion is localized to the site of the substitution. An oligoribonuceotide containing two deoxy residues directs specitic cleavage of RNA by E. coli RNase H. Structural requirements for cleavage are proposed for RNase V1 and RNase H.


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