Cleavage of a DNA–RNA–DNA/DNA chimeric substrate containing a single ribonucleotide at the DNA–RNA junction with prokaryotic RNases HII

• Published in: FEBS letters 2002-11, Vol.531 (2), p.204-208
• Main Authors: Haruki, Mitsuru, Tsunaka, Yasuo, Morikawa, Masaaki, Kanaya, Shigenori
• Format: Article
• Language: English
• Subjects: Bacillus subtilis; Bacillus subtilis - enzymology; Base Sequence; DNA - metabolism; DNA–RNA junction; DNA–RNA–DNA/DNA heteroduplex; Escherichia coli - enzymology; Kinetics; Nucleic Acid Heteroduplexes - chemistry; Nucleic Acid Heteroduplexes - metabolism; Ribonuclease H - metabolism; Ribonucleotides - metabolism; RNA - metabolism; Substrate Specificity; Thermococcus - enzymology; Thermococcus kodakaraensis; Type 2 ribonuclease H
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/S0014-5793(02)03503-2

We have analyzed the cleavage specificities of various prokaryotic Type 2 ribonucleases H (RNases H) on chimeric DNA–RNA–DNA/DNA substrates containing one to four ribonucleotides. RNases HII from Bacillus subtilis and Thermococcus kodakaraensis cleaved all of these substrates to produce a DNA segment with a 5′-monoribonucleotide. Consequently, these enzymes cleaved even the chimeric substrate containing a single ribonucleotide at the DNA–RNA junction (5′-side of the single ribonucleotide). In contrast, Escherichia coli RNase HI and B. subtilis RNase HIII did not cleave the chimeric substrate containing a single ribonucleotide. These results suggest that bacterial and archaeal RNases HII are involved in excision of a single ribonucleotide misincorporated into DNA.