Lesion specificity in the base excision repair enzyme hNeil1: modeling and dynamics studies

Abstract

Base excision repair (BER) is the major pathway employed to excise oxidized DNA lesions. Human Neil1, a versatile glycosylase in the BER pathway, repairs a diverse array of oxidative lesions; however, the most prevalent, 8-oxo-7,8-dihydroguanine (8-oxoG), is only weakly excised. The structural origin of hNeil1's ability to repair a variety of lesions but not 8-oxoG is a model system for connecting enzyme structure and lesion-recognition specificity. To elucidate structural properties determining hNeil1's substrate specificities, we have investigated it in complex with two pairs of representative well-repaired substrates: the R- and S-spiroiminodihydantoin (Sp) stereoisomers, nonplanar further oxidation products of guanine, and the 5R,6S- and 5S,6R-thymine glycol (Tg) stereoisomers, the most prevalent oxidative lesions of thymine. We also investigate the poorly repaired 8-oxoG. We employed molecular modeling and 10 ns molecular dynamics (MD) simulations. The results of our investigations provide structural explanations for the ability of hNeil1 to excise a variety of oxidative lesions: they possess common chemical features, namely, a pyrimidine-like ring and shared hydrogen bond donor-acceptor properties, which allow the lesions to fit well in the binding pocket, which is somewhat flexible. However, the planar 8-oxoG is not as well accommodated in the shallow and comparatively cramped recognition pocket; it has fewer hydrogen bonding interactions with the enzyme and a solvent exposed six-membered ring, consistent with its poor repair susceptibility by this enzyme.

Figure 1

(A) Structures of deoxyribonucleosides 8-oxoG, Sp R and S stereoisomers, and Tg 5R,6S and 5S,6R stereoisomers. (B) DNA sequences investigated and originally present in the bstFpg crystal structure. Glycosidic torsion angle χ is O4′ – C1′ – N9 – C4 for 8-oxoG and Sp, and O4′ – C1′ – N1 – C2 for Tg.

Figure 2

Plots of the all-atom root-mean-square deviations (RMSD) of the current relative to the starting structure as a function of time for the binding pocket (residues within 5 Å from any lesion atom), the whole complex, the helix αF region (residues 199 to 223), and the complex without this region. The αF region is distinctly more flexible than the rest of the complex.

Figure 3

Stereo views of (A) Electrostatic surface of hNeil1 with damaged DNA double strands (white and yellow) showing the positively charged DNA binding groove of hNeil1 (see text). Blue color is positive and red is negative. (B) DNA binding features of hNeil1 as described in text. The arginines are colored by atom with main chain and hydrogen atoms not shown. (C) DNA binding features of hNeil1 at the damaged site (see text). The lesion and protein residues are colored by atom. Hydrogen bonding interactions are marked with black lines. Damaged base and its base partner are in cyan. Structures from the last frame of the 10 ns simulations are presented. All stereo figures are prepared for viewing with a stereo viewer available at http://www.berezin.com/3D/viewers1.htm.

Figure 4

Stereo views of electrostatic surface (color code same as in Figure 3A) of the binding pocket of hNeil1 with Sp(syn) and Tg(anti) stereoisomers, and 8-oxoG(syn). Lesions are colored by atom. The sugar connected to the lesions is shown in yellow. Note the variable sizes and shapes of the binding pockets, as well as electrostatic interactions mainly indicating hydrogen bonds. Structures from the last frame of the 10 ns simulations are presented.

Figure 5

Stereo views of the binding pocket of hNeil1 with Tg and Sp stereoisomers, and 8-oxoG, highlighting hydrogen bonding interactions. See text for details. The enzyme is rendered in cartoon. Lesions and key residues are colored by atom. The sugar connected to the lesions is shown in yellow. Hydrogen bonding interactions are marked with black lines. Structures from the last frame of the 10 ns simulations are presented.

Figure 6

Hydrogen bonding interactions of hNeil1 residues Glu2 and Glu5 proposed for lesion recognition.