2’OH activation in RNA catalysis: metal-ion / nucleobase mechanisms in self-cleaving ribozymes

 

Zdeněk Chval1, Fabrice Leclerc2

 

1 Department of Laboratory Methods and Medical Technology, Faculty of Health and Social Studies, University of South Bohemia, J. Boreckého 27, 370 11 České Budějovice, Czech Republic

2 Laboratoire de Maturation des ARN et Enzymologie Moléculaire, Université Henri Poincaré, Faculté des Science, B.P. 239, Bd. des Aiguillettes, 54506 Vandoeuvre-lès-Nancy, France

chval@jcu.cz

 

The reaction catalyzed by a small catalytically active self-cleaving ribozymes is a phosphate ester hydrolysis that is thought to proceed via activation of the 2’OH group of the ribose moiety at the cleavage site. In this work the mechanism of the 2’OH activation is studied theoretically by DFT and MP2 methods. The effect of bulk solvent is treated with continuum solvation CPCM model. Mg2+ ion and the G-12 guanine nucleobase are considered as the catalysts of the proton transfer to activate 2’OH. Calculated free energy barriers of 2’OH activation are similar for Mg2+ ion and G-12. However negative charge on 2’O- is not stabilized enough by the bare G-12 nucleobase and further stabilization by other species (e.g. by metal ions) is necessary and/or the 2’-OH activation is concomitant with the nucleophilic attack on the phosphorus followed by an immediate cleavage reaction. Mg2+ ion stabilizes 2’O- by a direct coordination and kinetics of 2’OH activation by the Mg2+ ion is determined by the equilibrium between hexa- and penta-coordinated Mg2+ structures. The metal ion coordination to 2’OH lowers the pKa value of the 2’OH group by more than three log units.

 

This work is supported by the Czech Science Foundation (204/09/J010) and by the Ministry of Education of the Czech Republic (ME09062 KONTAKT).