APPLICATION OF HAMMERHEAD RIBOZYMES FOR STRUCTURAL STUDIES OF RIBOSOMAL 5S RNAS

We synthesized and used hammerhead ribozymes designed for the GUC and AUC target sequences located in the terminal loop C of animal and plan t 5S rRNAs to verify its involvement in a higher order structure forma tion of ribosomal RNA. The activity of both GUC and AUC specific riboz ymes were proven to be active against synthetic analogs of the stem II I and loop C in the two types of 5S rRNAs. In contrast we did not obse rve any ribozyme-catalysed hydrolysis within the loop C of the native 5S rRNAs. We considered two possible explanations of the results: an i nvolvement of the loop C in long range interactions and lack of a stru ctural flexibility to form proper tertiary structure of the complex wi th the ribozyme. From our data it seems that the single stranded nucle otides within the loop C can not form correct RNA duplex structure req uired for ribozyme activity. It is clear that hammerhead ribozymes can be used for inhibition of different steps of expression of genetic in formation only if the target sequence occurs in appropriate context, t hat allows to form correct secondary and tertiary structure of the rib ozyme-substrate complex.