Novel configurations of the hammerhead ribozyme: increased activity and reduced magnesium ion requirement.
The hammerhead ribozyme is one of the smallest of known catalytic RNAs. Its structure and mechanism have been examined using a broad range of approaches. Recently, crystal structures of the hammerhead have been reported. Within the crystal the hammerhead exists in a Y-shaped configuration in which helices I and II form the adjacent upper arms, while helix III forms the lower leg of the Y. Based on these findings, hammerheads in which helix I and II are constrained to remain adjacent and roughly parallel are expected to be catalytically active. We have examined this possibility with two novel hammerhead configurations, circular and circular/hairpin. These circular and circular/hairpin hammerhead ribozymes possess activity comparable to, and in some cases even greater than, non-constrained hammerheads. Since these novel ribozymes are constrained into a closed, and active conformation, they offer advantages for structure/function studies. Additionally, these novel ribozymes will be adventitious for antisense mediated gene inhibition, since they possess increased activity and a reduced requirement for magnesium ion, and are expected to be more resistant to nuclease degradation.
Wang
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Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah, 421 Wakara Way, Suite 318, Salt Lake City, Utah, 84108, USA.
Ruffner
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D002942:Circular Dichroism; D008274:Magnesium; D009690:Nucleic Acid Conformation; D016337:RNA, Catalytic / Q000737:chemistry*
D016337:RNA, Catalytic; D008274:Magnesium