Catalysis by RNase P RNA: Unique features and unprecedented active site plasticity

Tina Persson, Simona Cuzic, Roland K. Hartmann*

*Corresponding author for this work
47 Citations (Scopus)

Abstract

Metal ions are essential cofactors for precursor tRNA (ptRNA) processing by bacterial RNase P. The ribose 2′-OH at nucleotide (nt) -1 of ptRNAs is known to contribute to positioning of catalytic Me2+. To investigate the catalytic process, we used ptRNAs with single 2′-deoxy (2′-H), 2′-amino (2′-N), or 2′-fluoro (2′-F) modifications at the cleavage site (nt -1). 2′ modifications had small (2.4-7.7-fold) effects on ptRNA binding to E. coli RNase P RNA in the ground state, decreasing substrate affinity in the order 2′-OH > 2′-F > 2′-N > 2′-H. Effects on the rate of the chemical step (about 10-fold for 2′-F, almost 150-fold for 2′-H and 2′-N) were much stronger, and, except for the 2′-N modification, resembled strikingly those observed in the Tetrahymena ribozyme-catalyzed reaction at corresponding position. Mn2+ rescued cleavage of the 2′-N but also the 2′-H-modified ptRNA, arguing against a direct metal ion coordination at this location. Miscleavage between nt -1 and -2 was observed for the 2′-N-ptRNA at low pH (further influenced by the base identities at nt -1 and +73), suggesting repulsion of a catalytic metal ion due to protonation of the amino group. Effects caused by the 2′-N modification at nt -1 of the substrate allowed us to substantiate a mechanistic difference in phosphodiester hydrolysis catalyzed by Escherichia coli RNase P RNA and the Tetrahymena ribozyme: a metal ion binds next to the 2′ substituent at nt -1 in the reaction catalyzed by RNase P RNA, but not at the corresponding location in the Tetrahymena ribozyme reaction.

Original languageEnglish
JournalJournal of Biological Chemistry
Volume278
Issue number44
Pages (from-to)43394-43401
Number of pages8
ISSN0021-9258
DOIs
Publication statusPublished - 31.10.2003
Externally publishedYes

Research Areas and Centers

  • Academic Focus: Center for Infection and Inflammation Research (ZIEL)

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