STRUCTURAL-ANALYSIS OF THE P10 11-P12 RNA DOMAIN OF YEAST RNASE-P RNAAND ITS INTERACTION WITH MAGNESIUM/

The P10/11-P12 RNA domain of yeast RNase P contains several highly con served nucleotides within a conserved secondary structure. This RNA do main is essential for enzyme function in vivo, where it has a demonstr ated role in divalent cation utilization. To better understand the fun ction of this domain, its structure and alterations in response to mag nesium have been investigated in vitro. A secondary structure model of the P10/11-P12 RNA domain had been previously developed by phylogenet ic analysis. Computer modeling and energy minimization were applied to the Saccharomyces cerevisiae P10/11-P12 domain to explore alternative s and additional interactions not predicted by the phylogenetic consen sus. The working secondary structure models were challenged with data obtained from H-1 NMR and in vitro chemical and enzymatic probing expe riments. The solution structure of the isolated domain was found to co nform to the phylogenetic prediction within the context of the holoenz yme. Structure probing data also discriminated among additional base c ontacts predicted by energy minimization. The withdrawal of magnesium does not appear to cause gross refolding or rearrangement of the RNA d omain structure. Instead, subtle changes occur in the solution accessi bility of specific nucleotide positions, Most of the conserved nucleot ides reported to be involved in magnesium utilization in vivo also dis play magnesium-dependent changes in vitro.