PDBsum entry 1xc9

Transferase/DNA

PDB id: 1xc9

Contents

Protein chain

580 a.a.

DNA/RNA

Ligands

GLC-FRU

BAP

SO4 ×4

Metals

_MG

Waters ×373

References listed in PDB file

Key reference

Title

Structure of a high fidelity DNA polymerase bound to a benzo[a]pyrene adduct that blocks replication.

Authors

G.W.Hsu, X.Huang, N.P.Luneva, N.E.Geacintov, L.S.Beese.

Ref.

J Biol Chem, 2005, 280, 3764-3770. [DOI no: 10.1074/jbc.M411276200]

PubMed id

15548515

Abstract

Of the carcinogens to which humans are most frequently exposed, the polycyclic aromatic hydrocarbon benzo[a]pyrene (BP) is one of the most ubiquitous. BP is a byproduct of grilled foods and tobacco and fuel combustion and has long been linked to various human cancers, particularly lung and skin. BP is metabolized to diol epoxides that covalently modify DNA bases to form bulky adducts that block DNA synthesis by replicative or high fidelity DNA polymerases. Here we present the structure of a high fidelity polymerase from a thermostable strain of Bacillus stearothermophilus (Bacillus fragment) bound to the most common BP-derived N2-guanine adduct base-paired with cytosine. The BP adduct adopts a conformation that places the polycyclic BP moiety in the nascent DNA minor groove and is the first structure of a minor groove adduct bound to a polymerase. Orientation of the BP moiety into the nascent DNA minor groove results in extensive disruption to the interactions between the adducted DNA duplex and the polymerase. The disruptions revealed by the structure of Bacillus fragment bound to a BP adduct provide a molecular basis for rationalizing the potent blocking effect on replication exerted by BP adducts.

Figure 2.
FIG. 2. [BP]dG at the post-insertion site of the BF active site. Stereoview of the structure of BF bound to BP-modified DNA duplex (red) superimposed on the structure of BF bound to unmodified DNA duplex (gray). A schematic representing the BF active site is shown below.

Figure 3.
FIG. 3. [BP]dG-induced distortions to the BF polymerase active site. A, the C:[BP]dG base pair surrounded by omit electron density contoured at 3 with hydrogen bonds represented in black (dashed lines) accompanied by lengths. B, superposition of the BP-modified DNA duplex (red) and an unmodified DNA duplex (gray). The BP moiety of the [BP]dG protrudes into the DNA minor groove. C, comparison of BF protein side chain conformations when BF is bound to C:[BP]dG (red) and C:G (gray) at the post-insertion site (n-1 position). Hydrogen bonds are represented as dashed lines. D, model of BF in a ternary complex with [BP]dG (yellow) obstructing the insertion site to an incoming dCTP. A cognate base pair is shown in gray, and hydrogen bonds are represented by dashed lines.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2005, 280, 3764-3770) copyright 2005.

Secondary reference #1

Title

Observing translesion synthesis of an aromatic amine DNA adduct by a high-Fidelity DNA polymerase.

Authors

G.W.Hsu, J.R.Kiefer, D.Burnouf, O.J.Becherel, R.P.Fuchs, L.S.Beese.

Ref.

J Biol Chem, 2004, 279, 50280-50285. [DOI no: 10.1074/jbc.M409224200]

PubMed id

15385534

Note In the PDB file this reference is annotated as "TO BE PUBLISHED". The citation details given above were identified by an automated search of PubMed on title and author names, giving a percentage match of 95%.

Figure 1.
FIG. 1. A, chemical structures of G-AF and G-AAF. dR, deoxyribose ring. The proton and the N-acetyl group that differentiate G-AF from G-AAF are shown in orange. B, schematic of the polymerase active site. Sites through which the template base (red) traverses during replication are shown (see the Introduction for description).

Figure 4.
FIG. 4. Structure of BF with G-AF at the postinsertion site. A, structure of BF bound to G-AF at the postinsertion site (blue) superimposed with a structure of BF bound to an unmodified base at the postinsertion site (gray). G-AF (red) adopts an anti conformation and obstructs the n+1 template base from occupying the preinsertion site that is itself disordered. B, C:G-AF base pair surrounded by electron density contoured at 3.5 and calculated using Fourier coefficients (F[obs] - F[calc]) [calc] with C:G-AF omitted from the final model. Hydrogen bonds (dashed lines) are shown accompanied with bond lengths.

The above figures are reproduced from the cited reference with permission from the ASBMB

Secondary reference #2

Title

Structures of mismatch replication errors observed in a DNA polymerase.

Authors

S.J.Johnson, L.S.Beese.

Ref.

Cell, 2004, 116, 803-816. [DOI no: 10.1016/S0092-8674(04)00252-1]

PubMed id

15035983

Figure 2.
Figure 2. DNA Mismatches Bound at the Polymerase Postinsertion SiteThe bases are shown in the same orientation and location as the G•C base pair in Figure 1B. Left, hydrogen bonding pattern. Right, superimposition of the molecular surface of the mismatch (red) and cognate G•C base pair (yellow, PDB ID 1L3S) bound at the postinsertion site, highlighting differences in shape and location of the primer terminus.

Figure 4.
Figure 4. Extension of a G•T Mismatch by Successive Rounds of ReplicationThe conformation of the G•T mismatch is shown at each position (left), including interacting water molecules (red spheres). Dashed lines indicate potential hydrogen bonds. At the n-3 and n-4 positions, hydrogen bonds are shown between groups within the appropriate distance (≤3.2 Å) and correspond to tautomerization or ionization of one of the bases (see text). A schematic representation (right) of the mismatch complex, drawn and color coded as described in Figure 1, Figure 2 and Figure 3, indicates regions of the polymerase active site that are disrupted upon binding of the mismatch (red line). Mismatch binding at positions n-1 to n-4 along the DNA duplex binding region (gray) results in a distorted open conformation at the polymerase active site as described by mechanism 1 (Figure 3). The normal open conformation observed with homoduplexes is fully restored when the mismatch is bound at the n-6 position.

The above figures are reproduced from the cited reference with permission from Cell Press