4420
J. Phys. Chem. 1996, 100, 4420-4423
Ab Initio Molecular Orbital Study of the Structures of Purine Hydrates Anny-Odile Colson† and Michael D. Sevilla* Chemistry Department, Oakland UniVersity, Rochester, Michigan 48309 ReceiVed: October 12, 1995; In Final Form: December 1, 1995X
The structures of the isomers of purine hydrates [4(5)-hydroxy-5(4)-hydropurines] have been geometry optimized with ab initio quantum chemical methods at the 6-31G* basis set and with the semiempirical method PM3. These hydrates which can result from reduction of radical species formed by attack of hydroxyl radical at the 4,5 double bond in the purines, show significant geometrical distortion when compared to the natural bases. More specifically, the cis isomers adopt a “butterfly” conformation, while in the trans isomers, the pyrimidine and imidazole rings tilt opposite to each other. Our results predict the cis purine hydrate isomers are far more stable than the trans isomers by 10-18 kcal/mol at the 6-31G* level, whereas the 4-hydroxy5-hydropurines are found to be slightly more energetically stable than the 5-hydroxy-4-hydropurines. The “butterfly” conformation of the cis isomers constitutes a bulky lesion which will result in a significant distortion of the DNA helix.
Introduction Radiation-induced DNA damage has been widely recognized to occur in two principal ways formally referred to as the direct and indirect effect.1,2 Among the species resulting from either effect are the base hydroxyl adducts. These adducts not only result from hydroxide ion addition to the DNA base cation radicals formed from direct energy deposition on the DNA (direct effect) but also result from attack of hydroxyl radicals (one of the most damaging species formed upon low LET irradiation of the waters of hydration present in the vicinity of the DNA) to various sites of the natural DNA bases (indirect effect).1-4 Several of the DNA base hydroxyl adducts have been experimentally identified,1,5-7 and extensive work has focused on determining their redox properties,5,8-10 giving insight into their reactivities and their ability to induce strand break subsequent to hydrogen atom abstraction from the nearby sugar moieties. In a recent ab initio study performed on the most common hydroxyl and hydrogen adduct radicals of the four DNA bases,11 we have shown the trends in ionization potentials and electron affinities of these species are in good agreement with available experimental findings, and further complement these findings by proposing a consistent scheme of the relative redox properties of all radical intermediates. The stability of the various radical adducts has been shown to vary at pH 7,6 some undergoing dehydration to further give rise to yet other radicals, others undergoing ring opening. The fate of these radicals depends on their redox properties, those radicals with oxidizing properties interacting with reductants to form hydrates or to potentially regenerate the base by dehydration. Radicals with reducing properties interact with oxidants in kinetically controlled processes12 which may result in the formation of glycols. In addition to reacting with oxidizing or reducing species, these adducts can undergo radical-radical recombination. The formation of pyrimidine hydrates has been demonstrated in UV-irradiated solutions of free bases, nucleosides, and nucleotides as well as in a variety of double stranded polynucleotides.13-18 On the other hand, most of the hydroxyl adducts of the purines have been shown by pulse radiolysis † Present address: Department of Physiology and Biophysics, Mount Sinai School of Medicine, City University of New York, New York, New York 10029 X Abstract published in AdVance ACS Abstracts, February 1, 1996.
0022-3654/96/20100-4420$12.00/0
studies to dehydrate as radicals in aqueous solution.6,19-21 However, in DNA, reduction of the adducts is likely to result in the formation of purine hydrates. For example, the 8-hydroxyguanin-7-yl radical has been shown not to dehydrate in DNA but to undergo oxidation to 8-oxoguanine and reduction (after ring opening) to FAPYguanine.20 Thus for the 4,5 purine hydroxy adducts for which the ring opened precursor radical is not observed, formation of purine hydrates is very likely in DNA. Extensive theoretical calculations22 have focused on the energetics and structural preferences of the pyrimidine hydrates, contributing to the interpretation of experimental results on the biological consequences of pyrimidine lesions. As part of an ongoing theoretical investigation of the radiation-induced damage in DNA (for a review, see Colson and Sevilla23), in this work we investigate the energetics and geometrical features of the various possible purine hydrates and their potential implications on the conformation and function of DNA containing these lesions. Method of Calculation The 6-31G* polarization basis set24 and the PM3 semiempirical method25,26 implemented in the Gaussian 92 set of programs27 are used to fully geometry optimize the hydrates of the purines on Cray-C90 and IBM RS 6000 computers. The (x, y, z) coordinates are available in the supplementary material. Results and Discussion 1. Structures of Hydroxyhydropurines. The purine hydrates can exist in two different stereoisomeric forms: the cis isomers in which both the hydroxyl and hydrogen atom are bonded to the C4 and C5 sites on the same side of the plane of the base, and the trans isomer in which the OH and H groups bind to opposite sides of the plane. In this work, we have fully geometry optimized isomers of adenine and guanine, considering C4 and C5 as potential sites for hydroxyl attachment, resulting in the eight geometries shown in Figures 1 and 2. For the isolated bases studied here, only one of the enantiomer of each pair needs to be considered, as reflection through the (x,z) plane will result in identical structural features. However, in nucleosides and nucleotides, the sugar moiety and the DNA environ© 1996 American Chemical Society
Structures of Purine Hydrates
J. Phys. Chem., Vol. 100, No. 11, 1996 4421
Figure 1. 6-31G* geometry optimized cis and trans isomers of 4-hydroxy, 5-hydropurines.
ment will likely remove this degeneracy, and all enantiomeric pairs should be taken into consideration. One of the most striking structural features of the cis isomers is the significant bending of the bases in a “butterfly” shape around the C4-C5 axis. We have previously observed this characteristic in the hydroxyl adducts,11 but the attachment of the hydrogen moiety to the C4 (or C5) site further accentuates the bending of the structures, as can be seen from the angle measured between C8, the middle of the C4-C5 bond, and the middle of the N1-C2 bond presented in Table 1. The average decrease in this angle from that measured in the radical adducts is quite significant and amounts to ca. 26°. These data suggest that the increase in bending is independent of the nature of the base and the site of hydrogen attachment. Furthermore, a measure of the dihedral angles N3-C4-C5-C6 and N9-C4C5-N7 (Table 1) reveals both rings of each cis isomer remains nearly planar, as observed for the radical adducts. As shown in Figures 1 and 2, the conformation adopted by the trans purine hydrates significantly differs from that of the cis isomers. Indeed, in the trans isomers, the pyrimidine and imidazole rings of each base tilt in opposite directions around an axis perpendicular to the C4-C5 bond rather than bending around the C4C5 axis. This tilting appears to be quite symmetrical as shown by the near 180.0° values of the torsion angles N3-C4-C5N7 and C6-C5-C4-N9 (Table 1), suggesting comparable contributions from the hydroxyl and hydrogen moieties in distorting each individual base. In addition, the individual 5and 6-membered rings of the trans isomers deviate further from planarity than their counterparts in the cis isomers, as revealed by the dihedrals N3-C4-C5-C6 and N9-C4-C5-N7 (Table 1). For comparative purposes, the structures were also geometry optimized with the PM3 semiempirical method which has been shown to yield satisfactory bond distances and angles in many other systems.28 The structural features observed with this method are in excellent agreement with those observed with the 6-31G* basis set (Table 1). In addition to the structural changes observed in the C4-C5 region, the base pairing region appears to be mostly affected in the guanine hydrates if one expects the isolated DNA bases to be planar. Indeed, in adenine, C1, C6, N6, and H6 of both cis and trans isomers remain nearly coplanar, therefore preventing disruption of the hydrogen-bonding pattern with thymine, while in guanine the amino group hydrogens involved in one of the three hydrogen bonds to cytosine significantly protrude out of the plane. The deviation of the amino group hydrogen atoms of both purines from the plane of the base can be quantified by the selected angles presented in Table 2. It should be noted that recent theoretical investigations29 of the deformability of the amino group of guanine and adenine performed at the MP2/ 6-31G* basis set reveal the amino groups of both purines exhibit
Figure 2. 6-31G* geometry optimized cis and trans isomers of 5-hydroxy, 4-hydropurines.
significant sp3 pyramidalization. The angles calculated in this study29 are included in Table 2 and when compared to ours suggest that the nonplanarity of the amino group hydrogens observed upon hydrate formation does not significantly differ from that of the natural bases. In the natural bases the barrier to force an amino group coplanar with the ring is quite small (ca. 1 kcal/mol) and does not interfere with H-bond formation.29 High-level unconstrained geometry optimizations of the base pairs are needed to accurately predict the influence of the purine hydrates on the base pair geometries. It is interesting to note that in the structures presented in Figures 1 and 2, the hydroxyl group always points outwards except for the trans-4-hydroxy-5-hydroadenine. A single point calculation performed on this species in which the hydroxyl group was forced to point outwards as observed in the guanine counterpart resulted in a slightly less energetically stable structure (ca. 2 kcal/mol). This observation can be explained on the basis of atomic charges. In the trans-5-hydroxy species, the OH group points toward N7 which is slightly more negatively charged (-0.6) than N3 and N9 (-0.4). In the trans4-hydroxyguanine, N3 carries more negative charge (-0.6) than N7 and N9 (-0.4), while in adenine, N7 is the most negative of the three nitrogens (-0.6 vs -0.5(N3) and -0.3(N9)). These small differences in atomic charges appear to be sufficient to dictate the positioning of the OH group, although as described above, pointing of this moiety toward a slightly more positive nitrogen (i.e., N3 in the later adenine) is only responsible for a small energetic change in stability. Intermolecular interactions with the waters of hydration would likely overcome these small barriers. 2. Energetics of Hydroxyhydropurines. Total and relative energies of the cis and trans isomers of the purine hydrates investigated in this work at the 6-31G* and PM3 levels are presented in Table 3. Previous semiempirical calculations performed on the natural DNA bases at PM330 have shown this method yields heats of formation that are in reasonably good agreement with experiment. Hence, the results obtained in the present work with this method should contribute to verify the reliability of the results obtained at the 6-31G* level. The trends in relative stabilities calculated at both levels agree well with each other and suggest the cis isomers of the C4 and C5 hydroxypurine hydrates are more energetically stable than the trans isomers by an average 15 kcal/mol. This result is just what is expected from the structural features of the intermediate radicals described in a previous study.11 Indeed, we have shown the C4 and C5 radical precursors of the purine hydrates adopt
4422 J. Phys. Chem., Vol. 100, No. 11, 1996
Colson and Sevilla
TABLE 1: Selected Angles and Dihedral Angles Describing the Nonplanarity of the C4 and C5 Purine Adducts and Their Hydrates (degrees)a adenine 4-OH 5-H C8-(C4-C5)-(N1-C2) C6-C5-C4-N9c N3-C4-C5-N7c N3-C4-C5-C6d N9-C4-C5-N7d
5-OH 4-H
•A4(OH)
cis
trans
•A5(OH)
cis
trans
144 146.3 -135.6 -24.5 13.8
122 (120) 101.5 (112.8) -139.6 (-126.0) -19.4 (-6.8) -18.7 (-6.4)
-173.2 (-171.2) -164.0 (-164.9) 60.9 (59.7) -38.2 (-35.8)
153 143.8 -153.4 -29.0 19.4
119 (112) 142.0 (125.7) -95.2 (-114.0) 24.4 (4.7) 22.4 (7.0)
165.7 (167.6) 168.9 (167.0) -62.5 (-61.6) 37.2 (36.3)
G5(OH)
cis
trans
131 133.2 -126.6 -5.3 11.9
113 (118) 140.4 (131.3) -96.3 (-108.2) 22.0 (12.0) 22.0 (11.0)
164.7 (165.2) 164.2 (162.4) -66.3 (-66.3) 35.3 (33.9)
b
guanine 4-OH 5-H •
C8-(C4-C5)-(N1-C2) C6-C5-C4-N9c N3-C4-C5-N7c N3-C4-C5-C6d N9-C4-C5-N7d
b
5-OH 4-H •
G4(OH)
cis
trans
150 153.0 -133.5 32.5 -12.9
121 (117) 103.9 (114.8) -138.7 (-124.5) -16.8 (-4.7) -17.9 (-5.0)
-169.6 (-166.6) -160.6 (-160.6) 65.3 (65.7) -35.5 (-33.0)
a 6-31G* optimized angles. Values in parentheses are obtained with the PM3 semiempirical method. b Angle between C8 and center of 4,5 and 1,2 bonds; this is a measure of ring bending and is 180° in the planar parent DNA bases. c This angle is 180° in the parent DNA base.29 d This angle is 0° in the parent DNA base.29
TABLE 2: Angles and Dihedral Angles Describing the Nonplanarity of the Amino Groups of the Purine Hydrates (degrees)a,b adenine 4-OH 5-H Hb-N6-C6 H-N6-C6 Hb-N6-C6-N1 H-N6-C6-C5 a
guanine 5-OH 4-H
4-OH 5-H
5-OH 4-H
parent (ref 29)
cis
trans
cis
trans
parent (ref 29)
cis
trans
cis
trans
115.6 116.8 18.7 -21.1
117.6 118.5 -9.0 18.0
119.7 120.4 1.9 -4.0
118.7 120.3 2.9 -8.0
119.0 119.6 -6.0 15.4
115.4 111.1 38.7 -11.5
116.9 112.9 34.5 -10.7
116.5 113.0 38.6 -6.3
116.6 112.3 -37.2 10.7
114.9 111.6 -46.6 4.6
6-31G* optimized values. b Hb refers to the hydrogen atom involved in hydrogen bonding to the complementary pyrimidine in the base pairs.
a “butterfly” conformation, and since the cis isomers of the hydrates remain in a similar conformation, they are expected to be more stable than the trans isomers which adopt significantly different conformations as shown in Figures 1 and 2. In addition, Table 3 suggests the cis and trans isomers of the 4-hydroxy-5-hydropurines are energetically more stable than their 5-hydroxy-4-hydro counterparts. More specifically, the difference in energies between the trans isomers and the difference in energies between the cis isomers of adenine are nearly equal and amount to ca. 6 kcal/mol. It is interesting to note that this is only 2 kcal/mol lower than the difference in energy between the radicals •A(4)OH and •A(5)OH, suggesting that the structural changes and hence the energetics that accompany the deformations of each adduct induced upon reduction at the C4 or C5 sites of adenine are nearly identical. For guanine, •G(4)OH is more stable than •G(5)OH by nearly 8 kcal/mol. The same is true of trans-4-hydroxy-5-hydroguanine over trans-5-hydroxy-4-hydroguanine (i.e., ca. 7 kcal/mol), while the cis isomer of the 4-hydroxyguanine is only 1.3 kcal/ mol more stable than its 5-hydroxy counterpart. This result points to a greater stabilization of •G(5)OH vs •G(4)OH upon reduction of the radicals into the cis adducts. The greater stability of the 4-hydroxy-5-hydropurines over the 5-hydroxy-4-hydropurines can be explained on the basis of redox properties of their respective radical precursors. Indeed, in previous work,11 our ab initio calculations suggested the 4-hydroxypurine radicals are far more oxidizing than the 5-hydroxypurine radicals. It is therefore more likely for the former to be reduced and hence form hydrates which will be more energetically stable than the hydrates resulting from reduction of the less oxidizing 5-hydroxypurine radicals. Furthermore, we note that the calculated net atomic charges (at
TABLE 3: Energies (in kcal/mol) of the Purine Hydrates Relative to the Most Stable 4-OH Isomersa adenine 4-OH 5-H
guanine
5-OH 4-H
4-OH 5-H
5-OH 4-H
calcd level
cis
trans
cis
trans
cis
trans
cis
trans
6-31G* PM3
0 0
18.31 24.89
6.29 5.27
24.03 30.85
0 0
10.41 19.65
1.30 3.62
17.14 16.69
a All compounds are fully geometry optimized at each calculational level.
the 6-31G* basis set) at the radical site of the 4-hydroxy adducts (0.7) is far greater than that at the radical site of the 5-hydroxy adducts (0.3). This should greatly favor reduction of the 4-hydroxy adducts to the purine hydrates. Conclusion and Summary The present work on purine hydrates and our recent study on the C4 and C5 hydroxyl adducts of the purines foresees significant geometrical disruptions of the DNA helix upon formation of such lesions. Indeed, the “butterfly” geometry adopted by the radical adducts will facilitate the accessibility of the damaged site to reductive species. The resulting cis isomers of the purine hydrates will further disrupt the helix by their more pronounced bending around the C4-C5 axis while the tilted conformation adopted by the trans isomers may result in more subtle distortions of the biopolymer. Based on increased evidence that the bases can adopt nonplanar geometries in the amino groups,29,31,32 it appears that purine hydrates formation may not disrupt base pairing. The extent of the damage incurred will of course largely depend on the rates at which radical
Structures of Purine Hydrates formation, reduction, and dehydration occurs. The calculations presented in this work predict the 4-hydroxy-5-hydropurines are energetically favored over the 5-hydroxy-4-hydropurines, the cis isomers being more stable than the trans isomers by 10-18 kcal/mol. In previous work22,33 it was suggested that nonplanarity of saturated pyrimidine lesions is a likely cause of their differentiated biological effects. If this observation is valid for the purine hydrates studied in this work, our results suggest the 4- and 5-hydroxyhydropurines will result in different biological endpoints. At this time, there are no data on the structures of the 4- and 5-hydroxyhydropurines; thus our calculations provide a prediction of the structures and relative energies for these compounds. Additional experimental work on the stability and lifetime of these lesions in DNA is needed, as is additional theoretical work involving these lesions in molecular dynamics simulations of a larger DNA fragment. Presumably the large structural changes created by the bulkly cis isomers would produce changes in DNA conformation that should easily be detected by repair enzymes. Acknowledgment. We thank the National Cancer Institute of the National Institutes of Health (Grant RO1CA45424) and the Office of Health and Environmental Research of the Department of Energy (Grant DEFG0286ER60455) for support of this work. We thank the DOE National Energy Research Supercomputer Center at Lawrence Livermore National Laboratory for generous grants of computer time. Supporting Information Available: Coordinates (x, y, z) for species obtained at the 6-31G* and PM3 level (8 pages). This material is contained in many libraries on microfiche, immediately follows this article in the microfilm version of the journal, can be ordered from the ACS, and can be downloaded from the Internet; see any current masthead page for ordering information and Internet access instructions. References and Notes (1) Sevilla, M. D.; Becker, D. R. Soc. Chem. Spec. Period. Rep., Electron Spin Reson. 1994, 14, 130-165. (2) Becker, D.; Sevilla, M. D. AdV. Radiat. Biol., 1993, 17, 121-180.
J. Phys. Chem., Vol. 100, No. 11, 1996 4423 (3) von Sonntag, C. The Chemical Basis of Radiation Biology; Taylor and Francis: London, 1987. (4) Steenken, S. Chem. ReV. 1989, 89, 503. (5) Steenken, S. Free Radical Res. Commun. 1992, 16, 349. (6) O’Neill, P.; Fielden, E. M. AdV. Radiat. Biol. 1993, 17, 53-120. (7) Kuwabara, M. Radiat. Phys. Chem. 1991, 37, 691. (8) Vieira, A. J. S. C.; Steenken, S. J. Am. Chem. Soc. 1987, 109, 7441. (9) O’Neill, P. Radiat. Res. 1983, 96, 198. (10) Steenken, S. J. Chem. Soc., Faraday Trans. 1 1987, 83, 113. (11) Colson, A. O.; Sevilla, M. D. J. Phys. Chem. 1995, 99, 13033. (12) Fielden, E. M.; O’Neill, P.; Steenken, S. In The Early Effects of Radiation on DNA; NATO ASI Series H; Fielden, E. M., O’Neill, P., Eds.; Springer Verlag: Berlin, 1991. (13) Fisher, G. J.; Johns, H. E. Photochem. Photobiol. 1973, 18, 23. (14) Boorstein, R. J.; Cadet, J.; Hilbert, T.; Lustig, M.; O’Donnell, R.; Zuo, S.; Teebor, G. J. Chim. Phys. 1993, 90, 837. (15) Boorstein, R. J.; Hilbert, T. P.; Cadet, J.; Cunningham, R. P.; Teebor, G. W. Biochemistry 1989, 28, 6164. (16) Boorstein, R. J.; Hilbert, T. P.; Cunningham, R. P.; Teebor, G. W. Biochemistry 1990, 29, 10455. (17) Ganguly, T.; Weems, K. M.; Duker, N. J. Biochemistry 1990, 29, 7222. (18) Ganguly, T.; Duker, N. J. Nucleic Acids Res. 1991, 19, 3319. (19) Vieira, A. J. S. C.; Steenken, S. J. Am. Chem. Soc. 1990, 112, 6986. (20) Cadet, J.; Berger, M.; Buchko, G. W.; Joshi, P. C.; Morin, B.; Raoul, S.; Ravanat, J. L. In Radiation Damage in DNA, Structure/Function Relationships at Early Times; Fuciarelli, A. F., Zimbrick, J. D., Eds.; Battelle Press: Columbus OH, 1995. (21) Hankiewicz, E.; Bothe, E.; Schulte-Frohlinde, D. Free Radical Res. Commun. 1992, 16, 391. (22) Miaskiewicz, K.; Miller, J.; Osman, R. Biochim. Biophys. Acta 1994, 1218, 283. (23) Colson, A. O.; Sevilla, M. D. Int. J. Radiat. Biol. 1995, 67, 627. (24) Hariharan, P. C.; Pople, J. A. Chem. Phys. Lett. 1972, 16, 217. (25) Stewart, J. J. P. J. Comput. Chem. 1989, 10, 209. (26) Stewart, J. J. P. J. Comput. Chem. 1990, 11, 543. (27) Frisch, M. J.; Trucks, G. W.; Head-Gordon, M.; Gill, P. M. W.; Wong, M. W.; Foresman, J. B.; Johnson, B. G.; Schlegel, H. B.; Robb, M. A.; Replogle, E. S.; Gomperts, R.; Andres, J. L.; Raghavachari, K.; Binkley, J. S.; Gonzalez, C.; Martin, R. L.; Fox, D. J.; DeFrees, D. J.; Baker, J.; Stewart, J. J. P.; Pople, J. A. GAUSSIAN 92, ReVision B, C; Gaussian, Inc.: Pittsburgh, PA, 1992. (28) Levine, I. N. Quantum Chemistry; Prentice Hall: Englewood Cliffs, NJ, 1991. (29) Sponer, J. and Hobza, P. J. Phys. Chem. 1994, 98, 3161. (30) Hrouda, V.; Floria´n, J.; Hobza, P. J. Phys. Chem. 1993, 97, 1542. (31) Sponer, J.; Hobza, P. J. Mol. Struct. (THEOCHEM) 1994, 304, 35. (32) Riggs, N. V. Chem. Phys. Lett. 1991, 177, 447. (33) Miaskiewicz, K.; Miller, J.; Osman, R. Int. J. Radiat. Biol. 1993, 63, 677.
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