Unraveling the three-metal-ion catalytic mechanism of the DNA repair enzyme endonuclease IV

Departments of ^†Chemistry and Biochemistry and
^‖Pharmacology, and
^††Howard Hughes Medical Institute, University of California at San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0365;
^§Department of Molecular Biology and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, MB 4, La Jolla, CA 92037; and
^¶Life Sciences Division, Department of Molecular Biology, Lawrence Berkeley National Laboratory, Berkeley, CA 94720

Edited by Kenneth Merz, University of Florida, Gainesville, FL, and accepted by the Editorial Board December 1, 2006 (received for review April 28, 2006)

Abstract

Endonuclease IV belongs to a class of important apurinic/apyrimidinic endonucleases involved in DNA repair. Although a structure-based mechanistic hypothesis has been put forth for this enzyme, the detailed catalytic mechanism has remained unknown. Using thermodynamic integration in the context of ab initio quantum mechanics/molecular mechanics molecular dynamics, we examined certain aspects of the phosphodiester cleavage step in the mechanism. We found the reaction proceeded through a synchronous bimolecular (A_ND_N) mechanism with reaction free energy and barrier of −3.5 and 20.6 kcal/mol, in agreement with experimental estimates. In the course of the reaction the trinuclear active site of endonuclease IV underwent dramatic local conformational changes: shifts in the mode of coordination of both substrate and first-shell ligands. This qualitative finding supports the notion that structural rearrangements in the active sites of multinuclear enzymes are integral to biological function.

Footnotes

^‡To whom correspondence should be addressed. E-mail: iivanov{at}mccammon.ucsd.edu
Author contributions: I.I., J.A.T., and J.A.M. designed research; I.I. performed research; I.I. analyzed data; and I.I., J.A.T., and J.A.M. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS direct submission. K.M. is a guest editor invited by the Editorial Board.
This article contains supporting information online at www.pnas.org/cgi/content/full/0603468104/DC1.
Abbreviations:

QM,

quantum mechanics;

MM,

molecular mechanics;

endo IV,

endonuclease IV;

BER,

base excision repair;

AP,

apurinic/apyrimidinic;

ELF,

electron localization function.
Freely available online through the PNAS open access option.