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 o f approaches. Recently, crystal structures of the hammerhead have been reported. Within the crystal the hammerhead exists in a Y-shaped conf iguration in which helices I and II form the adjacent upper arms, whil e helix III forms the lower leg of the Y. Eased on these findings, ham merheads in which helix I and II are constrained to remain adjacent an d roughly parallel are expected to be catalytically active. We have ex amined this possibility with two novel hammerhead configurations, circ ular and circular/hairpin. These circular and circular/hairpin hammerh ead ribozymes possess activity comparable to, and in some cases even g reater than, non-constrained hammerheads. Since these novel ribozymes are constrained into a closed, and active conformation, they offer adv antages for structure/function studies. Additionally, these novel ribo zymes will be adventitious for antisense mediated gene inhibition, sin ce they possess increased activity and a reduced requirement for magne sium ion, and are expected to be more resistant to nuclease degradatio n. (C) 1998 Academic Press.