Crystal structure of a DNA decamer containing a cis-syn thymine dimer

Abstract

It is well known that exposure to UV induces DNA damage, which is the first step in mutagenesis and a major cause of skin cancer. Among a variety of photoproducts, cyclobutane-type pyrimidine photodimers (CPD) are the most abundant primary lesion. Despite its biological importance, the precise relationship between the structure and properties of DNA containing CPD has remained to be elucidated. Here, we report the free (unbound) crystal structure of duplex DNA containing a CPD lesion at a resolution of 2.0 A. Our crystal structure shows that the overall helical axis bends approximately 30 degrees toward the major groove and unwinds approximately 9 degrees, in remarkable agreement with some previous theoretical and experimental studies. There are also significant differences in local structure compared with standard B-DNA, including pinching of the minor groove at the 3' side of the CPD lesion, a severe change of the base pair parameter in the 5' side, and serious widening of both minor and major groves both 3' and 5' of the CPD. Overall, the structure of the damaged DNA differs from undamaged DNA to an extent that DNA repair proteins may recognize this conformation, and the various components of the replicational and transcriptional machinery may be interfered with due to the perturbed local and global structure.

Fig 1.

Structure of the cis-syn dimer (a) and numbering of the decamer sequence (b).

Fig 2.

Diagram showing the lattice packing of molecules in the unit cell. (a) One asymmetrical unit is composed of two duplex molecules (black–light gray or light gray–dark gray). They are held together by the stacking (b) and interstrand (c) hydrogen bonds. Two guanine bases are hydrogen bonded to each other in a symmetrical way. The bases involved are labeled.

Fig 3.

Multiple-wavelength anomalous diffraction electron density around the CPD site (1-s level). Selected bases are labeled. The figure was made by the program O (44).

Fig 4.

Schematic diagram illustrating the kink of the helix. Regular B-DNA and CPD-containing DNA are depicted in green and red, respectively. (a) Side view with a helical axis. (b) Top view.

Fig 5.

Stereo view representation of the detailed surroundings of the CPD site. Hydrogen bonds are drawn as dashed lines with distances. The view shows part of the major groove of the molecule. The hydrogen bond between N6H of A6 and O4 of T15 is significantly weakened. The phosphodeoxyribose backbone shows a sharply kinked or pinched structure. The figure was produced by using VMD (58).

Fig 6.

Electropotential surface models (a); (c) an ideal B-form decamer DNA (b); (d) crystal structure of the CPD containing decamer DNA (same sequence), with probe radii of 1.4 Å. The CPD site and corresponding site of the ideal B-form DNA are indicated by arrows. The duplex DNA molecule containing the CPD has an enlarged minor and major groove. The figure was produced by using VMD (58).

Fig 7.

Possible recognition mechanism of CPD site. (a) B-form DNA. (b) Model of the T4 endonuclease V enzyme bound to a CPD containing DNA. (c) Crystal structure of the T4 endonuclease V enzyme complexed with a CPD containing DNA (20). The direction of kinking (or bending) in the helical axis is the same between b and c.