Folding of the Hairpin Ribozyme in Its Natural Conformation Achieves Close Physical Proximity of the Loops

Abstract

The hairpin ribozyme is a self-cleaving motif found in the negative strand of the satellite RNA of some plant viruses. In its natural context, the ribozyme comprises four helices, two of which contain conserved formally unpaired loops, that are adjacent arms of a four-way RNA junction. We show that the arms that would carry these loops are brought close together in the global conformation of the isolated junction. Using fluorescence resonance energy transfer, we demonstrate a two-magnesium ion-dependent conformational transition of the complete ribozyme that brings the loop-bearing arms into close physical proximity. The ribozyme is active as a four-way junction, and the rate of cleavage may be modulated by the conformation of the four-way junction.