Transition-state stabilization in Escherichia coli ribonuclease P RNA-mediated cleavage of model substrates. Issue 1 (3rd October 2013)

• Record Type: Journal Article
• Authors: Wu, Shiying; Chen, Yu; Mao, Guanzhong; Trobro, Stefan; Kwiatkowski, Marek; Kirsebom, Leif A.
• Is Part Of: Nucleic acids research. Volume 42:Issue 1(2014)
• Page Start: 631
• Page End: 642
• Publication Date: 2013-10-03
• Subjects: Nucleic acids -- Periodicals; Molecular biology -- Periodicals; 572.805
• DOI: 10.1093/nar/gkt853
• Languages: English
• ISSNs: 0305-1048

Abstract

Abstract: We have used model substrates carrying modified nucleotides at the site immediately 5′ of the canonical RNase P cleavage site, the −1 position, to study Escherichia coli RNase P RNA-mediated cleavage. We show that the nucleobase at −1 is not essential but its presence and identity contribute to efficiency, fidelity of cleavage and stabilization of the transition state. When U or C is present at −1, the carbonyl oxygen at C2 on the nucleobase contributes to transition-state stabilization, and thus acts as a positive determinant. For substrates with purines at −1, an exocyclic amine at C2 on the nucleobase promotes cleavage at an alternative site and it has a negative impact on cleavage at the canonical site. We also provide new insights into the interaction between E. coli RNase P RNA and the −1 residue in the substrate. Our findings will be discussed using a model where bacterial RNase P cleavage proceeds through a conformational-assisted mechanism that positions the metal(II)-activated H2 O for an in-line attack on the phosphorous atom that leads to breakage of the phosphodiester bond.