Transition-state stabilization in Escherichia coli ribonuclease P RNA-mediated cleavage of model substrates
2014 (English)In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 42, no 1, 631-642 p.Article in journal (Refereed) Published
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 H2O for an in-line attack on the phosphorous atom that leads to breakage of the phosphodiester bond.
Place, publisher, year, edition, pages
2014. Vol. 42, no 1, 631-642 p.
IdentifiersURN: urn:nbn:se:uu:diva-221063DOI: 10.1093/nar/gkt853ISI: 000331136000056OAI: oai:DiVA.org:uu-221063DiVA: diva2:708158