The OsGEN-L protein from Oryza sativa possesses Holliday junction resolvase activity as well as 5'-flap endonuclease activity

• Published in: Journal of biochemistry (Tokyo) 2012-03, Vol.151 (3), p.317-327
• Main Authors: Yang, Yantao, Ishino, Sonoko, Yamagami, Takeshi, Kumamaru, Toshihiro, Satoh, Hikaru, Ishino, Yoshizumi
• Format: Article
• Language: English
• Subjects: DNA Repair - genetics; DNA Repair - physiology; DNA, Cruciform - metabolism; Flap Endonucleases - genetics; Flap Endonucleases - metabolism; Holliday Junction Resolvases - genetics; Holliday Junction Resolvases - metabolism; Oryza - enzymology; Oryza sativa; Plant Proteins - genetics; Plant Proteins - metabolism; Proliferating Cell Nuclear Antigen - metabolism
• ISSN: 0021-924X; 1756-2651
• DOI: 10.1093/jb/mvr145

OsGEN-L has a 5'-flap endonuclease activity and plays an essential role in rice microspore development. The Class 4 RAD2/XPG family nucleases, including OsGEN-L, were recently found to have resolving activity for the Holliday junction (HJ), the intermediate of DNA strand recombination. In this study, we performed a detailed characterization of OsGEN-L, as a structure-specific endonuclease. Highly purified OsGEN-L was prepared as the full-length protein for in vitro endonuclease assays using various structured DNAs, and the 5'-flap endonuclease activity, which is stimulated in a PCNA-dependent manner, was demonstrated. In addition, the in vitro HJ resolving activity of OsGEN-L represents the first such activity originating from plant cells. OsGEN-L cleaved HJ at symmetrically related sites of the branch point. However, the two branched strands seemed to be cleaved individually, and not cooperatively, by each OsGEN-L monomer protein. The substrate specificity suggests that OsGEN-L functions in multiple processes of DNA metabolism in rice cells.