From the Department of Chemistry, Wesleyan University, Middletown, Connecticut 06459
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ABSTRACT |
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Natural and exogenous processes can give rise to abasic sites with either a purine or pyrimidine as the base on the opposing strand. The solution state structures of the apyrimidinic DNA duplex, with D6 indicating an abasic site,
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INTRODUCTION |
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Damage to DNA bases can arise from a number of routes including oxidative stress, the action of various chemical agents, and by radiative processes (1-5). Common examples of base damages include the spontaneous deamination of cytosine to uracil, the oxidation of thymine to thymine glycol or urea, and the photochemical production of thymine dimer adducts. The first step in repair of damaged DNA in vivo is often the hydrolytic cleavage of the C-N bond between the sugar and the damaged or unusual base to generate an aldehydic abasic site that is sometimes referred to as an apurinic/apyrimidinic or AP1 site (1-5).
The role of the base opposite the abasic site is of interest because there are many routes by which an abasic site can be generated (2, 3, 6, 7). Deamination of a dC residue followed by the action of N-uracil DNA glycosylase leads to dG opposite the abasic site. Oxidation of dT to thymine glycol or urea and the subsequent action of a glycosylase lead to dA opposite an abasic site, whereas base damage to dA or dG can lead to having dC or dT opposite the abasic site. The base opposite the abasic site may have structural, dynamic, or other properties that affect the chemical reactivity of the duplex DNA, its recognition by proteins, or its interactions in subsequent repair reactions (8, 9). Also, it now appears that some polymerases may place all four bases opposite an abasic site and not only dA residues (10) leading to both apurinic and apyrimidinic sites. The structure of damaged DNA is sequence-dependent as is the structure of undamaged DNA, and the presence of an abasic site within a curved DNA sequence can have large, long range structural effects (11). Results on the structures of repair enzymes have led to the suggestion that the abasic site can be recognized directly (12).
Damaged DNA plays roles in controlling the cell cycle, and these roles are of considerable and growing interest because of their importance in apoptosis and carcinogenesis as noted in recent reviews (13-18). It now appears that there are "checkpoints" for the presence of damaged DNA which need to be passed before a cell can go from G1 to G2 (18). The processes by which these checkpoints control the G1 to G2 passage are not yet known. It is also not known how many types of damaged DNA are recognized at these checkpoints nor how many different procedures are used to recognize damaged DNA (19-22). Normal cells with damaged DNA are programmed to undergo apoptosis, and apparently only cells with damaged DNA and which avoid this step can become transformed into tumor cells (2, 16, 18, 23-26). Abasic sites are also poisons for topoisomerase, which may be important in apoptosis (27-29).
In many cells the predominant pathway to the formation of aldehydic abasic sites is the spontaneous deamination of cytosine to uracil (5, 6, 30-34). The enzyme N-uracil glycoslyase excises uracil to form an aldehydic abasic site as shown in Scheme I.
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For a typical Escherichia coli there are about 40-400 such events per cell division, and in a typical mammalian cell 4,000-40,000 uracil are formed per cell division (2, 35, 36). The number of damaged sites in a particular genome depends on the state of the cell, and some damaged sites may not be repaired until the G1 to G2 passage occurs. The number of abasic sites in a "typical" human cell is not known because the rates of damage and repair are dependent on many factors. Ames and co-workers (37) have estimated that there are more than 10,000 damaged sites, of all types, per typical human cell at any given time. The presence of a direct relationship between DNA damage and cancer is known, but the number and types of damage to DNA which are required for transformation to occur are just now being determined.
The cleavage of the glycosidic bonds is catalyzed by DNA glycosylases,
which were first identified by Lindahl and Nyberg in 1974 (38), and
many distinct classes of glycosylases are now known (6). Crystal
structures of two uracil glycosylases have been determined (39, 40) as
have the structure of uracil glycosylase inhibitor protein free and
complexed to uracil glycosylase (41-43). The abasic site is not a
chemically unique species but is an equilibrium mixture (44-47) of
- (I), and
- (II), hemiacetals that are
2-deoxy-D-erythro-pentofuranoses, of aldehyde
(III), and of hydrated aldehyde (IV), as depicted below. The
3'-cleavage of the abasic site catalyzed by AP lyases is via a
syn
-elimination reaction (6) (Scheme
II).
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Subsequent to the formation of the aldehydic abasic site the repair
process can continue with the cleavage of both the 5'- and
3'-phosphodiester bonds. The 3'-cleavage reaction catalyzed by UV
endonuclease V of bacteriophage T4 and by E. coli
endonuclease III or the enzyme formamidopyrimidine-DNA glycosylase is a
-elimination that proceeds with syn stereochemistry, with
abstraction of the 2'-pro-S proton, as discussed elsewhere
(44-48). The 5'-cleavage is via a delta elimination when catalyzed by
formamidopyrimidine-DNA glycosylase (40). After the abasic site is
cleaved at both the 3'- and 5'-phosphodiester linkages the site is
repaired via synthesis and ligation of the DNA (7).
There have been a number of investigations of the effects of unrepaired aldehydic abasic sites on replication. Studies on the effect of abasic sites on both in vivo and in vitro replication have been carried out on a variety of DNA polymerases (2, 5, 8, 49-51). Recent results indicate that DNA damage can affect DNA replication at sites remote from the site of damage (52) indicative of interactions of the polymerases with the DNA at positions remote from the site of damage.
The presence of an unrepaired aldehydic abasic site can lead to a stable mutation as well as having effects on transcription (2, 53, 54). The results to date suggest that the presence of an abasic site slows down but does not block transcription (53-55). The base most commonly placed at the position complementary to the abasic site in transcription is rA. This is the same preference as found for some, but not all, DNA polymerases (10, 23, 56).
To determine the details of the structure of a DNA duplex containing an abasic site we have investigated the apyrimidinic duplex,
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MATERIALS AND METHODS |
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NMR Procedures-- NMR data were acquired at 500 MHz on a Varian Inova and at 400 MHz on a Varian Unityplus spectrometer, at Wesleyan, and at 500 MHz on a Bruker 500 MHz DMX spectrometer at the University of Wisconsin, Madison, using methods described previously (11, 57-59) and modified as discussed below. All 31P experiments were obtained at 161 MHz using the 400-MHz Varian Unityplus. All of the Varian NMR data were processed using VNMR software, and all of the Bruker data were processed using Felix 95.0 software. All of the Varian two-dimensional data were obtained using States-Haberkorn and all of the Bruker data using TPPI.
NOESY experiments on AD were run on the Bruker 500 at Madison with the sample in 2H2O at 25 °C with mixing times of 100 and 200 ms and a 1.6-s equilibration delay. The spectral width in each dimension was 5,000 Hz. 512 t1 increments were acquired with 64 transients/t1 point and 1024 complex points in the t2 dimension. The data were processed with a sinebell apodization in both dimensions before 2048 × 2048 Fourier transformation. These spectra were used for NOESY cross-peak quantification. NOESY experiments on CD were run on the Varian 500 with the sample in 2H2O at 15 °C with mixing times of 100 and 250 ms and a 1-s equilibration delay. The spectral width in each dimension was 6,000 Hz. 448 t1 increments were acquired with 48 transients/t1 point and 4096 complex points in the t2 dimension. The data were processed with a Gaussian weighting in both dimensions before 4096 × 1024 Fourier transformation. These spectra were used for NOESY cross-peak quantification. NOESY watergate experiments on CD were run on the Varian 500 with the sample in 90% H2O and 10% 2H2O at 15 and at 5 °C with a mixing time of 100 ms using a 1-s equilibration delay. Spectral widths in both dimensions were 12,000 Hz, and processing with Gaussian weighting in both dimensions before a 4096 × 1024 Fourier transformation was used. ROESY experiments, with watergate suppression of the water resonance, on CD in 90% H2O and 10% 2H2O, were run using the Varian 500 with the sample at 15, 5, and 1 °C with a mixing time of 50 ms and a 1-s equilibration delay. The spectral width was 12,000 Hz in both dimensions, and the data were transformed using Gaussian shifted weighting functions into 4096 × 1024 real points. Quiet-NOESY experiments (60) on CD and AD were run on the Varian 400 with the sample in 2H2O at 15 °C with a mixing time of 250 ms and a 1-s equilibration delay. In the middle of the mixing time a 180° shaped Gaussian pulse was applied on the aromatic region in one experiment and on the methyl region in a separate Quiet-NOESY experiment. The spectral width in each dimension was 5,000 Hz. 256 t1 increments were acquired with 48 transients/t1 point and 4096 complex points in the t2 dimension. The data were processed using Gaussian apodization in both dimensions before 4096 × 1024 Fourier transformation. Band-selective TOCSY experiments (61) were run on the Varian 400 with the samples in 2H2O and at 15 °C for CD and 27 °C for AD with mixing times of 70 ms and a 1-s equilibration delay. Two 180° Gaussian shaped pulses were applied to the H3' region during the spin echo before the TOCSY spin lock. The spectral width in each dimension was 5,000 Hz. 300 t1 increments were acquired with 48 transients/t1 increment and 4096 complex points in the t2 dimension. The J scale for coupling to 31P was set to 2 and to 3 in separate experiments. The data were processed with a Gaussian apodization in both dimensions before 4096 × 1024 Fourier transformation. PECOSY spectra were acquired for the AD and CD samples at 400 MHz with 31P decoupling during the evolution period. The data were collected as 2,000 complex points in t2 and 512 complex points in t1. The F2 spectral width was 3,200 Hz, and the F1 spectral width was 2,600 Hz. 256 transients were acquired for each t1 increment. A Gaussian weighting function was used in both dimensions, and the data set was zero filled to 2048 × 2048 real points. TOCSY experiments were run on AD and CD with a mixing time of 80 ms with the samples in 2H2O at 400 MHz. The water resonance was presaturated during the 1.5-s equilibrium delay. The spectral width in both dimensions was 6,000 Hz. 2,048 complex points were collected in the t2 dimension and 256 complex points in t1. Gaussian weighting was applied before Fourier transformation into 2048 × 1024 real points. A series of one-dimensional inversion recovery experiments was run on AD and CD to allow determination of the chemical shifts of the adenine H2 resonances on the basis of their relatively long T1 values. These spectra were acquired with the samples in 2H2O at 400 MHz. An equilibrium delay of 12 s was used, and the inversion recovery delay time was arrayed from 0.3 to 2.0 s in equal steps. The Bruker 500 MHz spectrometer was used to obtain a 125-ms ROESY experiment on AD in 2H2O with the sample at 25 °C. A spectral width of 6,000 Hz was used in each dimension. The data were collected as 2,000 complex t1 points and 512 complex t2 points. Gaussian weighting was applied in both dimensions, and the data were zero filled to 2,000 × 2,000 real points.Quantitation and Assignment of NOE Cross-peaks--
The spectra
were assigned using the procedures described previously for abasic
site-containing DNAs (11, 58, 62). The standard sequential
connectivities of B-form DNA could be used except at the site of the
damage. In the form the H1' is spatially closer to the H2" than the
H2'. In the
form the H1' is spatially closer to the H2' than the
H2". AD NOE cross-peak volumes were quantified from the data obtained
at 500 MHz at mixing times of 100 and 200 ms using FELIX 95.0 software.
The NOE cross-peak volumes, of CD, were quantified in the data obtained
at 500 MHz with mixing times of 100 and 250 ms using VNMR software. For
each assigned cross-peak the volume over a standard area was
determined.
Structure Determination Procedures--
Structure refinement by
restrained molecular dynamics using X-PLOR 3.1 was performed as
described previously (11, 57-59, 63) with the following modifications.
The optimum weighting of the NOE constraints was found to be 60 kcal/mol, and the experimental volumes were used as the constraints.
For AD, at each mixing time, 233 NOE volume constraints were used for
the and for the
structure, and the refinements of the two
structures were carried out independently. For CD, at each mixing time,
258 NOE volume constraints were used for the two
structures, and
the refinements of the two structures were carried out independently.
During each structure calculation 34 dihedral constraints for the
ribose (H1'-C1'-C2'-H2'-and H1'-C1'-C2'-H2'') were used as well as 20 dihedral constraints for the backbone (P-O3'-C3'-H3') with a weighting
of 40 kcal/mol.
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RESULTS AND DISCUSSION |
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The one-dimensional spectra of the imino and exchangeable protons and of the phosphorus nuclei of the two damaged DNA duplexes are shown in Fig. 1. The spectra of the two samples are quite distinct in each of these three regions. The spectra of the nonexchangeable protons indicate that many protons have differing chemical shifts depending on whether the residue opposite the abasic site is a dA or dC. The imino region of the two spectra are the most similar of the three regions shown. In both cases the net number of imino protons integrates to about 10, indicating that all of the base pairs are present at 5°. The chemical shifts of the imino protons of the AT base pairs are to lower field for AD than for CD, and there is slightly better dispersion of the GC imino chemical shifts for the AD duplex than for the CD duplex.
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The 31P chemical shifts from the sites adjacent to the damaged site are different in AD and CD. The CD duplex has two signals that are significantly downfield of the main group, whereas the AD duplex has two that are only somewhat downfield of the main group. The most downfield 31P resonance in CD is from the phosphorus between residues 6 and 7, and the next most downfield one is from the phosphorus between residues 5 and 6. The two downfield resonances from AD are from the phosphorus sites between residues 6 and 7 and between 5 and 6, but there was not sufficient spectral resolution to determine which was the one slightly more downfield.
Thus, the one-dimensional data indicate that both damaged DNAs form duplexes, generally in the B-family with all possible base pairs present. However, the CD sample appears to exhibit the larger perturbation from B-form near the damaged site. The overall patterns of the chemical shifts are similar in both cases, but a sufficient number of differences suggests that there are significant structural differences resulting from whether dC or dA is opposite the abasic site.
The proton assignments of AD have been presented previously (58), and
many of these are indicated in Fig. 2.
The same assignment strategy was used for CD and led to the conclusion
that there are two forms present rather than equal amount of
and
. The two
forms will be referred to as
N3 and
O2
because of the basis of their main structural difference. We have
previously examined the AD duplex and a similar sequence context with
both dA and dG opposite the abasic site as well as an abasic site in a
curved DNA context, and essentially equal amounts of
and
had
been observed in each of these cases (11, 58, 62).
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The assignments of most of the CD resonances could be made using the
standard interresidue and intraresidue NOEs supplemented by information
from TOCSY and PE/DQCOSY experiments as discussed above. However, no
appreciable signals from an abasic site in the form could be
detected from the CD sample at 15 °C. At higher temperatures small
amounts of an
form could be detected, but these spectra were not
analyzed in detail because under these conditions signals from three
forms of CD were present. The results of NOESY experiments allowed
clear identification of
and
forms on the basis of the
H1'-H2'/H2" NOEs. In the
form the H1' is spatially closer to the
H2' than the H1', and the H1'-H2' NOE is larger than the H1'-H2". In
the
form the H1' has a larger scalar coupling to the H2" than the
H1'. The converse is the case for the
form. The lack of an
form
of CD is consistent with results obtained quite some time ago on a
7-mer duplex with a dC opposite the abasic site, which indicated that
essentially only one anomeric form was present based on results on a
sample with the 1'C labeled with 13C (44).
After the NOE volumes were quantified and the scalar couplings determined, this information was used to determine the structures of the two forms of each damaged DNA via restrained molecular dynamics. The predicted, back-calculated NOE results for each of the forms were then combined and compared with the experimental results as shown in Figs. 2 and 3. It is seen that the agreement between the experimental NOEs and those predicted by the structures of the DNAs is quite good in the regions shown and is equally good in the regions not shown. The main difference between the structure determination methods used here and those used previously is the inclusion of a water molecule at the damaged site during the restrained molecular dynamics. Our earlier results had shown that there is likely to be hydrogen bonding involving a water molecule at abasic sites in duplex DNAs (58), and the inclusion of the water molecule during the trajectories gave rise to structures that were in better agreement with the experimental results than could be obtained without the inclusion of the water molecules into the simulations.
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The resulting structures of the AD duplex with the and
forms of
the abasic site are shown in Fig. 4. The
overlay of five structures, obtained at 2-ps intervals, is shown at the
top. The overlays show that the trajectories are stable over
the 10-ps time interval with the r.m.s. deviation for the
structures at 0.48 Å and 0.4 Å for the
structures over this time
period. The trajectories appeared to become stable at, or before, about
100 ps. The structures shown at the bottom of Fig. 4 are the
average structure over the 10-ps period, and the expansion of the
region near the damaged site shows the DNA backbone, including the
sugar, in a thin line, the bases in a thicker
line, and the water molecule in CPK mode. This representation
emphasizes that in the
conformation a water molecule can be well
situated for hydrogen bonding to both the abasic site 1'OH as well as
to the hydrogen bond donor N3 of dA17. This is essentially
what was observed when the structure was determined and the position of
the water molecule examined afterward rather than being included in the
structure determination (58). It appears that the presence of the water
molecule allows the
form of AD to adopt a structure that is quite
similar to that of B-form DNA at the damaged site.
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The structure of the form of AD is shown in Fig. 4. The optimum
position of the water molecule is not as well suited for hydrogen
bonding to the abasic site nor the dA in the position opposite the
abasic site as in the
form. The position of the dA17 is
much closer to the abasic site in the
form than is the case for the
form, and this is part of the structural differences between the
two forms.
The structural determination of the CD sample followed much the same
route. The structures of the two forms were determined independently,
the structures were each used to back-calculate results for the two
structural forms, and these results were combined and compared with the
experimental results as shown in Fig. 3. The variation for the CD case
is that both structural forms have the abasic site in the form. In
one of the two structures the water molecule hydrogen bonds to the N3
of dC17, referred to as
N3, and in the other the water
molecule hydrogen bonds to the O2 of dC17, referred to as
O2. The overlays of the structures at 2-ps time steps over a 10-ps
time interval for both of these structures are shown in Fig.
5. The two structural forms are in slow
exchange, on the NMR time scale, as indicated by the presence of
distinct cross-peaks, for the two structural forms, which are separated
by 10-20 Hz for the dT16 H6 to Me intraresidue cross-peak, as shown in Fig. 6. No cross-peaks that
could be attributed to exchange between the two forms were observed in
the NOESY, ROESY, or TOCSY data sets.
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The overlays of the structures show that the trajectories are stable
over the 10-ps time interval with the r.m.s. deviation for the N3
structures at 0.56 Å and 0.55 Å for the
O2 structures over this
time period. As shown at the bottom of Fig. 5 the prime difference between the two structures is the position of
dC17 with the water molecule within hydrogen bonding
distance of the 1'OH of the abasic site and with either the O2 or N3 of
dC17.
Analysis of the original set of CD trajectories indicated that the
experimental results are only consistent with the presence of two
forms, both with a abasic sites, and that the dC17
residue was the site of conformational heterogeneity. Additional
preliminary trajectories carried out without constraints on the
position of the water molecule did not converge because the position of
the water was found to have transitions between positions that allowed hydrogen bonding to the N3 or O2 of dC17. Because NOE and
ROE connectivities between water and residues near the damaged site could be observed, the positions of the water molecules, in the two
forms, were constrained as described above. Cross-peaks between water
and the two forms of the H2 of dA7, the H2 of
dA5, and the amino protons of dC17 were
observed in NOESY spectra as well as in the ROESY spectrum shown in
Fig. 6. These experimentally observed water-DNA contacts are consistent
with the positions of the water molecules found in the trajectories of
the
N3 and
O2 forms.
The structures of AD and CD were used as a basis to model the surface
accessibilities of the residues of these damaged DNAs. As we have shown
previously, the accessible surface area appears to be a good surrogate
for "extrahelicity" as well as indicating which portions of the
damaged DNAs are available for recognition without alteration in the
structure (11, 57). The surface accessibilities of most of the residues
are the same in the cases of AD and CD with the exceptions of
dT18 and dC19 as shown in Fig.
7. The residue dT18 has a
significantly higher surface accessibility in both forms of CD than it
does in either form of AD. The residue dC19 has the highest
surface accessibility in the N3 form of CD.
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An overall view of the surface accessibilities of the damaged DNAs is
given in Fig. 8 for the case of a 1.5 Å probe molecule. The surface accessibilities of the and
forms of
AD are shown, on the left, with the abasic site coded in
red, the dA17 opposite the abasic site in
blue, the dA5 and dA7 adjacent to
the abasic site in yellow, and the dT16 and
dT18 coded in green. The positions of major
differences in surface accessibility are dA17, which is
much more accessible from the major groove in the
form than in the
, and dA5 and dA7, which are more accessible
from the minor groove in the
form than in the
. The surface
accessibilities of CD are shown on the right side of Fig. 8
with
N3 on the top and
O2 on the bottom.
The color coding is the same as for AD with the exception that it is
dC17 in blue.
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Thus, the structures of the two forms of CD are distinct from those of
AD near the damaged site, and the extent of distortion from B-form DNA
is more pronounced for CD. The least distorted structure is that of AD
with the form of the abasic site. The
form of the abasic site
appears to be able to hydrogen bond to a water molecule that can also
hydrogen bond to the N1 of dA17 on the opposing strand.
This positioning of this water molecule allows the abasic site and
dA17 to adopt positions analogous to those of B-form
DNA.
When the abasic site is in the form this hydrogen bonding
arrangement apparently cannot be adopted. When the 1'OH of the abasic
site is in the "down" position, associated with the
form, the
arrangement of the abasic site, the water molecule, and
dA17 is less favorable than in the
form in which the
water molecule can effectively bridge the two strands. Thus, the most
favorable arrangement that occurs in the AD
case is when the water
molecule hydrogen bonds to both the 1'OH hydrogen of the abasic site
and the N1 of dA17. In neither of these structural forms is
the abasic site itself significantly exposed, consistent with the
chemical stability of the abasic site in duplex DNA.
The origin of the structural differences between the CD and AD forms
appears to be that dC is simply too small to hydrogen bond to a water
molecule that is simultaneously hydrogen bonded to the abasic site
while the dC is near the position it would occupy in B-form DNA. Thus,
the structure near the abasic site is more distorted when dC is present
than dA as evidenced both by the larger accessible surface and the
absence of an form in CD. The structures of the
O2 and
N3
forms have a water molecule hydrogen bonded to the dC residue. In the
actual solution state there will be many water molecules interacting
with the damaged sites of both AD and CD, and only the position of a
single water molecule has been considered here.
The structure of a duplex DNA with a thymine glycol opposite dA in the same sequence context has also been determined (57). This structure is shown along with that of AD at the top of Fig. 9. The overall features of the AD and thymine glycol duplex DNAs are relatively similar near the damaged site in the undamaged strand. However, the thymine glycol residue has considerable extrahelicity, and the backbone is more distorted in the damaged strand of the thymine glycol DNA than is the case for AD.
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The structure of a duplex DNA with an abasic site opposite dA has been
determined in a curved dA tract DNA context (11), and the structures of
dA and the dA tract-damaged DNA are shown in Fig. 9. The comparison
shows that the structures of the two damaged DNAs are strikingly
distinct. The structures of the and the
forms are considerably
different in the two sequence contexts. This comparison shows that the
structure of a DNA containing an abasic site is not determined solely
by the type of damage but by the sequence context in which the damage
appears.
It may have been the case that the structure of duplex DNA containing an abasic site is determined primarily by the "hole" left by the excised base. However, the results presented here indicate that the structure is determined, in part, by the base on the opposing strand, because the structures with dA and dC opposite the abasic site have distinct structural features. Comparison of the structural results on AD and on the abasic site in the curved context shows that the sequence context in which the damage occurs also has a role in determining the structure of damaged DNA. Comparison of the structures of damaged DNAs with the same sequence but with different types of damage, abasic site or thymine glycol, shows that the structure is dependent on the type of damage.
The surface accessibilities of all of these structures have been determined. In each case the accessibilities of at least some of the residues at and near the damaged site are considerably greater than those of undamaged DNAs. The structures of these damaged DNAs also all have alterations in the width and regularity of both the minor and major grooves as well as the ratio of the widths of the two grooves. None of these forms of damage appears to induce a large change in the radius of the DNAs at or near the damaged site.
The structures of only a small number of the naturally occurring damaged DNA sites have been determined. In addition to the structures discussed above the structures and other properties of duplex DNAs with 8-oxoguanine have been examined (65-68). Although some generalizations may be suggested by this small number of structures it is expected that a somewhat larger set of structures needs to be determined and the biological properties of the set determined. This allows a detailed examination of which structural features of damaged DNAs can be recognized as "generic" damaged DNA and which can be recognized as being associated with a specific type of damaged DNA.
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FOOTNOTES |
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* This research was supported in part by American Cancer Society Grant NP-750. The 400-MHz NMR spectrometer was purchased with support from National Science Foundation Grant BIR 93-03077. The 500-MHz spectrometer was purchased with support from National Science Foundation Grant BIR-95-12478 and by the Camille and Henry Dreyfus Foundation.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
The atomic coordinates (1a9i and 1a9j for the and
forms
of the AD duplex, respectively, and 1a9g and 1a9h for the
N3 and
O2 forms of CD, respectively) have been deposited in the Protein
Data Bank, Brookhaven National Laboratory, Upton, NY.
To whom correspondence should be addressed. Tel.: 860-685-2668;
Fax: 860-685-2211; E-mail: pbolton{at}wesleyan.edu.
1 The abbreviations used are: AP, apurinic/apyrimidinic; AD, apyrimidinic duplex; CD, apurinic duplex; NOESY, nuclear Overhauser effect spectroscopy; ROESY, rotating frame Overhauser effect spectroscopy; TOCSY, total correlation spectroscopy; PECOSY, phased easy correlation spectroscopy; NOE, nuclear Overhauser effect; r.m.s., root mean square; DQCOSY, double quantum filter correlation spectroscopy.
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REFERENCES |
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