Sequence-specific cleavage of RNA by a hybrid ribonuclease H.
Site-specific cleavage of the 22-, 132- and 534-base RNAs by the DNA/protein hybrid RNase H were examined. The 22-base RNA was chemically synthesized, and 132- and 534-base RNAs were prepared by run-off transcription. The hybrid enzyme cleaves these RNAs, which contain a single target sequence, primarily at the unique phosphodiester bond within the target sequence. The hybrid enzyme performs multiple turnovers, and at a substrate/enzyme ratio of 10:1 the RNAs are almost completely cleaved by the hybrid enzyme at 37 degrees C within 1 h. We propose that hybrid RNase H molecules with various oligodeoxyribonucleotides function as RNA restriction enzymes and are useful for structural and functional studies of RNA.
Nakai
C
C
Protein Engineering Research Institute, Osaka, Japan.
Konishi
A
A
Komatsu
Y
Y
Inoue
H
H
Ohtsuka
E
E
Kanaya
S
S
D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D006868:Hydrolysis; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016297:Mutagenesis, Site-Directed; D009693:Nucleic Acid Hybridization; D012313:RNA / Q000737:chemistry / Q000378:metabolism*; D016914:Ribonuclease H / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity
D012313:RNA; D004247:DNA; D016914:Ribonuclease H