IQUIR   05412
INSTITUTO DE QUIMICA ROSARIO
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
STUDY OF THE MICRO RNA PRECURSOR PRO-CESSING MECHANISM BY QM/MM COMPUTATIONAL SIMULATIONS
Autor/es:
MORENO, DIEGO M.; PUZZOLO, JUAN L.; DRUSIN, SALVADOR IVÁN; RASIA, RODOLFO
Reunión:
Congreso; Reunión conjunta de las Sociedades de Biociencias; 2017
Resumen:
Rnase III is the main protein in the processing of microRNA precursors (pri-miRNA) in bacteria. It is composed by two domains: dsRBD, which recognizes and binds the target precursors, and RIIID, which processes the pri-miRNA. The processing by RIIID is performed through a hydrolysis reaction, in which there is a nucleophilic attack on the phosphate group of the RNA backbone by an activated H2O molecule or a OH- ion previously coordinated to a Mg2+ ion of the active site. The proposed mechanism on this reaction is based on structural studies that are not able to capture the thermodynamics of the process, nor the protonation states of the residues and solvent molecules involved in the mechanism with atomistic detail. In this regard, computational simulation techniques can provide useful and detailed information about the reaction mechanism.To study the hydrolysis mechanism through hybrid quantum mechanics/molecular mechanics (QM/MM) methods, we performed 50 ns of classical molecular dynamics (MD) simulation on the initial system. Next, we carried out 30 QM/MM steered molecular dynamics (SMD) simulations from different structures taken from the previous MD, in which we forced the nucleophile to attack the phosphate group. The irreversible work obtained in each SMD was used to calculate the free energy profile of the reaction, through the Jarzynksi equality.Our results show that the the nucleophile is an OH- ion and not a H2O molecule, as the former's energy barrier is considerably lower. Additionally, the network of hydrogen bridges around the active site is important for its regeneration after the reaction. On the other hand, by mutating key residues on the active site which bind the Mg2+ ion, we observed that its role is not only activating the nucleophile, but arranging and stabilizing the active site by neutralizing the negative charge of the phosphate, nucleophile, and protein residues.Keywords: microRNA, QM/MM, hydrolysis mechanism.