PMID:19917291
| Citation |
Sun, L, Campbell, FE, Yandek, LE and Harris, ME (2010) Binding of C5 protein to P RNA enhances the rate constant for catalysis for P RNA processing of pre-tRNAs lacking a consensus (+ 1)/C(+ 72) pair. J. Mol. Biol. 395:1019-37 |
|---|---|
| Abstract |
The RNA subunit of the ribonucleoprotein enzyme ribonuclease P (RNase P (P RNA) contains the active site, but binding of Escherichia coli RNase P protein (C5) to P RNA increases the rate constant for catalysis for certain pre-tRNA substrates up to 1000-fold. Structure-swapping experiments between a substrate that is cleaved slowly by P RNA alone (pre-tRNA(f-met605)) and one that is cleaved quickly (pre-tRNA(met608)) pinpoint the characteristic C(+1)/A(+72) base pair of initiator tRNA(f-met) as the sole determinant of slow RNA-alone catalysis. Unlike other substrate modifications that slow RNA-alone catalysis, the presence of a C(+1)/A(+72) base pair reduces the rate constant for processing at both correct and miscleavage sites, indicating an indirect but nonetheless important role in catalysis. Analysis of the Mg(2)(+) dependence of apparent catalytic rate constants for pre-tRNA(met608) and a pre-tRNA(met608) (+1)C/(+72)A mutant provides evidence that C5 promotes rate enhancement primarily by compensating for the decrease in the affinity of metal ions important for catalysis engendered by the presence of the CA pair. Together, these results support and extend current models for RNase P substrate recognition in which contacts involving the conserved (+1)G/C(+72) pair of tRNA stabilize functional metal ion binding. Additionally, these observations suggest that C5 protein has evolved to compensate for tRNA variation at positions important for binding to P RNA, allowing for tRNA specialization. |
| Links |
PubMed PMC3410724 Online version:10.1016/j.jmb.2009.11.027 |
| Keywords |
Base Pairing; Base Sequence; Catalysis; Consensus Sequence; Escherichia coli/genetics; Escherichia coli/metabolism; Escherichia coli Proteins/chemistry; Escherichia coli Proteins/genetics; Escherichia coli Proteins/metabolism; Kinetics; Magnesium/metabolism; Models, Molecular; Molecular Sequence Data; Nucleic Acid Conformation; Protein Conformation; RNA Precursors/chemistry; RNA Precursors/genetics; RNA Precursors/metabolism; RNA Processing, Post-Transcriptional; RNA, Bacterial/chemistry; RNA, Bacterial/genetics; RNA, Bacterial/metabolism; Ribonuclease P/chemistry; Ribonuclease P/genetics; Ribonuclease P/metabolism; Substrate Specificity; Thermodynamics |
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