Dataset Information

Quantum Mechanical/Molecular Mechanical Study of the HDV Ribozyme: Impact of the Catalytic Metal Ion on the Mechanism.

ABSTRACT: A recent crystal structure of the precleaved HDV ribozyme along with biochemical data support a mechanism for phosphodiester bond self-cleavage in which C75 acts as a general acid and bound Mg(2+) ion acts as a Lewis acid. Herein this precleaved crystal structure is used as the basis for quantum mechanical/molecular mechanical calculations. These calculations indicate that the self-cleavage reaction is concerted with a phosphorane-like transition state when a divalent ion, Mg(2+) or Ca(2+), is bound at the catalytic site but is sequential with a phosphorane intermediate when a monovalent ion, such as Na(+), is at this site. Electrostatic potential calculations suggest that the divalent metal ion at the catalytic site lowers the pK(a) of C75, leading to the concerted mechanism in which the proton is partially transferred to the leaving group in the phosphorane-like transition state. These observations are consistent with experimental data, including pK(a) measurements, reaction kinetics, and proton inventories with divalent and monovalent ions.

SUBMITTER: Ganguly A

PROVIDER: S-EPMC3233192 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature

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Publications

Quantum Mechanical/Molecular Mechanical Study of the HDV Ribozyme: Impact of the Catalytic Metal Ion on the Mechanism.

Ganguly Abir A Bevilacqua Philip C PC Hammes-Schiffer Sharon S

The journal of physical chemistry letters 20111101 22

A recent crystal structure of the precleaved HDV ribozyme along with biochemical data support a mechanism for phosphodiester bond self-cleavage in which C75 acts as a general acid and bound Mg(2+) ion acts as a Lewis acid. Herein this precleaved crystal structure is used as the basis for quantum mechanical/molecular mechanical calculations. These calculations indicate that the self-cleavage reaction is concerted with a phosphorane-like transition state when a divalent ion, Mg(2+) or Ca(2+), is b ...[more]

PMID: 22163069

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Project description:DNA polymerases play a crucial role in the cell cycle due to their involvement in genome replication and repair. Understanding the reaction mechanism by which these polymerases carry out their function can provide insights into these processes. Recently, the crystal structures of human DNA polymerase lambda (Pollambda) have been reported both for pre- and post-catalytic complexes [García-Díaz et al., DNA Repair 3 (2007), 1333]. Here we employ the pre-catalytic complex as a starting structure for the determination of the catalytic mechanism of Pollambda using ab initio quantum mechanical/molecular mechanical methods. The reaction path has been calculated using Mg(2+) and Mn(2+) as the catalytic metals. In both cases the reaction proceeds through a two-step mechanism where the 3'-OH of the primer sugar ring is deprotonated by one of the conserved Asp residues (D490) in the active site before the incorporation of the nucleotide to the nascent DNA chain. A significant charge transfer is observed between both metals and some residues in the active site as the reaction proceeds. The optimized reactant and product structures agree with the reported crystal structures. In addition, the calculated reaction barriers for both metals are close to experimentally estimated barriers. Energy decomposition analysis to explain individual residue contributions suggests that several amino acids surrounding the active site are important for catalysis. Some of these residues, including R420, R488 and E529, have been implicated in catalysis by previous mutagenesis experiments on the homologous residues on Polbeta. Furthermore, Pollambda residues R420 and E529 found to be important from the energy decomposition analysis, are homologous to residues R183 and E295 in Polbeta, both of which are linked to cancer. In addition, residues R386, E391, K422 and K472 appear to have an important role in catalysis and could be a potential target for mutagenesis experiments. There is partial conservation of these residues across the Pol X family of DNA polymerases.

Dataset Information

Quantum Mechanical/Molecular Mechanical Study of the HDV Ribozyme: Impact of the Catalytic Metal Ion on the Mechanism.

Publications

Quantum Mechanical/Molecular Mechanical Study of the HDV Ribozyme: Impact of the Catalytic Metal Ion on the Mechanism.

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