Hydrolytic endonucleolytic ribozyme (HYER) is programmable for sequence-specific DNA cleavage.

Science (New York, N.Y.) 2024 Feb 02

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Ribozymes are catalytic RNAs with diverse functions including self-splicing and polymerization. This work aims to discover natural ribozymes that behave as hydrolytic and sequence-specific DNA endonucleases, which could be repurposed as DNA manipulation tools. Focused on bacterial group II-C introns, we found that many systems without intron-encoded protein propagate multiple copies in their resident genomes. These introns, named HYdrolytic Endonucleolytic Ribozymes (HYERs), cleaved RNA, single-stranded DNA, bubbled double-stranded DNA (dsDNA), and plasmids in vitro. HYER1 generated dsDNA breaks in the mammalian genome. Cryo-electron microscopy analysis revealed a homodimer structure for HYER1, where each monomer contains a Mg2+-dependent hydrolysis pocket and captures DNA complementary to the target recognition site (TRS). Rational designs including TRS extension, recruiting sequence insertion, and heterodimerization yielded engineered HYERs showing improved specificity and flexibility for DNA manipulation.

Citation

Zi-Xian Liu, Shouyue Zhang, Han-Zhou Zhu, Zhi-Hang Chen, Yun Yang, Long-Qi Li, Yuan Lei, Yun Liu, Dan-Yuan Li, Ao Sun, Cheng-Ping Li, Shun-Qing Tan, Gao-Li Wang, Jie-Yi Shen, Shuai Jin, Caixia Gao, Jun-Jie Gogo Liu. Hydrolytic endonucleolytic ribozyme (HYER) is programmable for sequence-specific DNA cleavage. Science (New York, N.Y.). 2024 Feb 02;383(6682):eadh4859