title: Stereospecific formation of the (R)-gamma-hydroxytrimethylene interstrand N2-dG:N2-dG cross-link arising from the gamma-OH-1,N2-propano-2'-deoxyguanosine adduct in the 5'-CpG-3' DNA sequence.
abstract: Acrolein reacts with dG to form hydroxylated 1,N(2)-propanodeoxyguanosine (OH-PdG) adducts. Most abundant are the epimeric 3-(2-deoxy-beta-D-erythro-pentofuranosyl)-5,6,7,8-tetrahydro-8-hydroxypyrimido[1,2a] purin-10(3H)-ones, commonly referred to as the gamma-OH-PdG adducts. When placed complementary to deoxycytosine in duplex DNA, these undergo rearrangement to the N(2)-(3-oxopropyl)-dG aldehyde. The latter forms diastereomeric interstrand N(2)-dG:N(2)-dG cross-links in the 5'-CpG-3' sequence. Here we report the structure of the stereochemically favored (R)-gamma-hydroxytrimethylene N(2)-dG:N(2)-dG interstrand DNA cross-link in 5'-d(G(1)C(2)T(3)A(4)G(5)C(6)X(7)A(8)G(9)T(10)C(11)C(12))-3' x 5'-d(G(13)G(14)A(15)C(16)T(17)C(18)Y(19)C(20)T(21)A(22)G(23)C(24))-3' (X(7) is the dG linked to the alpha-carbon of the carbinolamine linkage, and Y(19) is the dG linked to the gamma-carbon of the carbinolamine linkage; the cross-link is in the 5'-CpG-3' sequence). The structure was characterized using isotope-edited (15)N nuclear Overhauser enhancement spectroscopy heteronuclear single quantum correlation (NOESY-HSQC) NMR, in which the exocyclic amines at X(7) or Y(19) were (15)N-labeled. Analyses of NOE intensities involving Y(19) N(2)H indicated that the (R)-gamma-hydroxytrimethylene linkage was the major cross-link species, constituting 80-90% of the cross-link. The X(7) and Y(19) imino resonances were observed at 65 degrees C. Additionally, for the 5'-neighbor base pair G(5) x C(20), the G(5) imino resonance remained sharp at 55 degrees C but broadened at 65 degrees C. In contrast, for the 3'-neighbor A(8) x T(17) base pair, the T(17) imino resonance was severely broadened at 55 degrees C. Structural refinement using NOE distance restraints obtained from isotope-edited (15)N NOESY-HSQC data indicated that the (R)-gamma-hydroxytrimethylene linkage maintained the C(6) x Y(19) and X(7) x C(18) base pairs with minimal structural perturbations. The (R)-gamma-hydroxytrimethylene linkage was located in the minor groove. The X(7) N(2) and Y(19) N(2) atoms were in the gauche conformation with respect to the linkage, which maintained Watson-Crick hydrogen bonding of the cross-linked base pairs. The anti conformation of the hydroxyl group with respect to C(alpha) of the tether minimized steric interaction and, more importantly, allowed the formation of a hydrogen bond between the hydroxyl group and C(20) O(2) located in the 5'-neighboring base pair G(5) x C(20). The formation of this hydrogen bond may, in part, explain the thermal stability of this carbinolamine interstrand cross-link and the stereochemical preference for the (R) configuration of the cross-link.
authors.affiliation: Department of Chemistry and Center in Molecular Toxicology, Vanderbilt University, Nashville, Tennessee 37235, USA.
authors.forename: Hai
authors.initials: H
authors.lastname: Huang
authors.forename: Hye-Young
authors.initials: HY
authors.lastname: Kim
authors.forename: Ivan D
authors.initials: ID
authors.lastname: Kozekov
authors.forename: Young-Jin
authors.initials: YJ
authors.lastname: Cho
authors.forename: Hao
authors.initials: H
authors.lastname: Wang
authors.forename: Albena
authors.initials: A
authors.lastname: Kozekova
authors.forename: Thomas M
authors.initials: TM
authors.lastname: Harris
authors.forename: Carmelo J
authors.initials: CJ
authors.lastname: Rizzo
authors.forename: Michael P
authors.initials: MP
authors.lastname: Stone
mesh_terms: D020029:Base Pairing; D018899:CpG Islands; D003432:Cross-Linking Reagents / Q000737:chemistry*; D003521:Cyclopropanes / Q000737:chemistry*; D004247:DNA / Q000737:chemistry*; D018736:DNA Adducts / Q000737:chemistry*; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000737:chemistry; D006860:Hydrogen Bonding; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D009690:Nucleic Acid Conformation; D013237:Stereoisomerism; D013816:Thermodynamics
chemical_list: D003432:Cross-Linking Reagents; D003521:Cyclopropanes; D018736:DNA Adducts; C424742:OH-1,-N(2)-propano-2'-deoxyguanosine; D004247:DNA; C030797:cyclopropane; D003849:Deoxyguanosine