Structural analysis of the P10/11-P12 RNA domain of yeast RNase P RNA and its interaction with magnesium.
The P10/11-P12 RNA domain of yeast RNase P contains several highly conserved nucleotides within a conserved secondary structure. This RNA domain is essential for enzyme function in vivo, where it has a demonstrated role in divalent cation utilization. To better understand the function of this domain, its structure and alterations in response to magnesium have been investigated in vitro. A secondary structure model of the P10/11-P12 RNA domain had been previously developed by phylogenetic analysis. Computer modeling and energy minimization were applied to the Saccharomyces cerevisiae P10/11-P12 domain to explore alternatives and additional interactions not predicted by the phylogenetic consensus. The working secondary structure models were challenged with data obtained from 1H NMR and in vitro chemical and enzymatic probing experiments. The solution structure of the isolated domain was found to conform to the phylogenetic prediction within the context of the holoenzyme. Structure probing data also discriminated among additional base contacts predicted by energy minimization. The withdrawal of magnesium does not appear to cause gross refolding or rearrangement of the RNA domain structure. Instead, subtle changes occur in the solution accessibility of specific nucleotide positions. Most of the conserved nucleotides reported to be involved in magnesium utilization in vivo also display magnesium-dependent changes in vitro.
Ziehler
W A
WA
Department of Biological Chemistry, University of Michigan, Ann Arbor 48109-0606, USA.
Yang
J
J
Kurochkin
A V
AV
Sandusky
P O
PO
Zuiderweg
E R
ER
Engelke
D R
DR
D001483:Base Sequence; D003198:Computer Simulation; D017124:Conserved Sequence; D017931:DNA Primers / Q000235:genetics; D004722:Endoribonucleases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D008274:Magnesium / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D016337:RNA, Catalytic / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D012331:RNA, Fungal / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D043262:Ribonuclease P; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D013816:Thermodynamics
D017931:DNA Primers; D016337:RNA, Catalytic; D012331:RNA, Fungal; D004722:Endoribonucleases; D043262:Ribonuclease P; D008274:Magnesium