Structure–activity relationship of pseudoknot-type hammerhead ribozyme reveals key structural elements for enhanced catalytic activity^†

Abstract

The pseudoknot-type hammerhead ribozyme (PK-HHRz) is known to be activated by a pseudoknot interaction between loops I and II. To obtain maximal activation through the pseudoknot formation, we studied the structure–activity relationship of PK-HHRz. From these studies, the structural requirements of the PK-HHRz cleavage reaction were clearly defined. In addition, we discovered a PK-HHRz with higher cleavage activity than the wild-type sequence. Although modifications generally disrupt the activity of enzymes, in this case the elongation of loop II increased the activity of PK-HHRz. These new findings will form a structural basis for designing PK-HHRz variants for gene-therapeutic/manipulating agents and biochemical/nanotechnological tools.

Keywords:

• Hammerhead ribozyme
• pseudoknot-type hammerhead ribozyme
• RNA

Additional information

Funding

This study was supported by Platform for Drug Discovery, Informatics, and Structural Life Science from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan. This study was also supported by grant-in-aid for Scientific Research (C) (19K05705 to Y.T.), (B) (24310163 to Y.T.) and (C) (20550145 to Y.T.) from MEXT, Japan; Human Frontier Science Program (Young Investigator Grant to Y.T) from the Human Frontier Science Program Organization, France. M.Y. is the recipient of a Japan Society for the Promotion of Science (JSPS) Research Fellowship for Young Scientists. grant-in-aid for Scientific Research (B), MEXT, Japan; Platform for Drug Discovery, Informatics, and Structural Life Science from MEXT, Japan; grant-in-aid for Scientific Research (C), MEXT, Japan.