Single-molecule studies of group II intron ribozymes

Abstract

Group II intron ribozymes fold into their native structure by a unique stepwise process that involves an initial slow compaction followed by fast formation of the native state in a Mg²⁺-dependent manner. Single-molecule fluorescence reveals three distinct on-pathway conformations in dynamic equilibrium connected by relatively small activation barriers. From a most stable near-native state, the unobserved catalytically active conformer is reached. This most compact conformer occurs only transiently above 20 mM Mg²⁺ and is stabilized by substrate binding, which together explain the slow cleavage of the ribozyme. Structural dynamics increase with increasing Mg²⁺ concentrations, enabling the enzyme to reach its active state.