Skip to main content
NCBI home page
NIHPA Author Manuscripts logoLink to NIHPA Author Manuscripts
. Author manuscript; available in PMC: 2006 Oct 12.
Published in final edited form as: Chem Res Toxicol. 2006 Jan;19(1):16–19. doi: 10.1021/tx050250z

The 5’-GNC Site for DNA Interstrand Cross-Linking is Conserved for Diepoxybutane Stereoisomers

Julie T Millard †,*, Trevor C Hanly †, Kris Murphy ‡, Natalia Tretyakova ‡,*
PMCID: PMC1599837  NIHMSID: NIHMS12527  PMID: 16411651

Abstract

The bifunctional alkylating agent 1,2,3,4-diepoxybutane forms interstrand DNA-DNA cross-links between the N7 positions of deoxyguanosine residues on opposite strands of the duplex. For racemic diepoxybutane, these cross-links predominate within 5’-GNC/3’CNG sequences, where N is any nucleotide. We used denaturing polyacrylamide gel electrophoresis (dPAGE) to examine the role of stereochemistry in the cross-linking reaction, subjecting a restriction fragment to cross-linking with S,S DEB, R,R DEB, or meso DEB. DNA cross-links generated by each isomer were isolated by dPAGE, and the sites of cross-linking were identified by sequencing gel analysis of DNA fragments generated by hot piperidine cleavage. We found that the 5’-GNC consensus sequence of racemic DEB is conserved, but the efficiencies of cross-linking vary, with S,S > R,R > meso DEB. These results help explain the observed differences between the biological activities of DEB stereoisomers.

INTRODUCTION

1,2,3,4-Diepoxybutane (DEB) is a cytotoxic bifunctional alkylating agent that produces mutations and chromosomal aberrations (1). As an active metabolite of 1,3-butadiene, DEB has been implicated in the high incidence of leukemia among workers in the styrene-butadiene industry (2-5). On the other hand, S,S DEB is believed to be the active metabolite of the pro-drug treosulfan (L-threitol 1,4 bismethanesulphonate), used in the treatment of advanced ovarian cancer (6).

Because DEB is about two orders of magnitude more mutagenic and cytotoxic than the monoepoxide metabolites of butadiene, its biological activity has been attributed to the formation of interstrand cross-links within DNA (7, 8). The isolation of N7-linked guanine-guanine conjugates from DEB-treated DNA (9) originally led to the proposal of interstrand cross-linking at 5’-GC sites, which have the minimal N7-to-N7 distance within canonical B-DNA (10). The structural assignment of the DEB-DNA conjugate as 1,4-bis(guan-7-yl)-2,3-butanediol was later confirmed independently (11). However, the preferential site of cross-linking was found to be at 5’-GNC sequences (where N is any base) for racemic DEB within both DNA oligomers and defined sequence nucleosomal core particles (12, 13). This core sequence preference is further modulated by the secondary influence of flanking sequences (14).

Most prior studies of DNA cross-linking have been carried out with racemic DEB, although some early reports suggested that the three DEB stereisomers (Chart 1) differ in their biological activity. S,S DEB is the most genotoxic and cytotoxic, followed by R,R and then meso (15-17). Recently, the different stereoisomers were reported to have distinct reactivities with DNA (18). While meso DEB forms comparable amounts of interstrand and intrastrand cross-links, R,R DEB induces predominantly interstrand adducts with smaller amounts of instrastrand bis-N7-guanine lesions, and S,S DEB selectively gives rise to interstrand DNA-DNA lesions (18). These findings are consistent with prior reports that meso DEB is a less effective interstrand cross-linker than either S,S or R,R DEB (15, 19).

Chart 1.

Chart 1

Open in a new tab

Structures of DEB stereoisomers: R,R DEB (1a), S,S DEB (1b), and meso DEB (1c).

In this study, we examined the sequence preferences of the three optical isomers of DEB within a 152 base pair (bp) restriction fragment. We found that while the three compounds vary in their efficiencies of forming interstrand cross-links, all stereoisomers share the 5’-GNC consensus site for cross-linking.

EXPERIMENTAL PROCEDURES

Caution: DEB is mutagenic and a suspect human carcinogen and must be handled appropriately. Cross-Linking Reactions. Racemic DEB was purchased from Sigma-Aldrich Chemical Company (Milwaukee, WI). Optically active (S,S and R,R) and meso DEB were prepared as described previously (18). A 152 bp EcoRI-RsaI restriction fragment from the plasmid pXP-11 (20), containing a portion of the Xenopus borealis 5S RNA gene and 5’-flanking region, was 3’-end radiolabeled and purified as described previously (21). Cross-linking was initiated by addition of DEB (1 μL of either racemic; S,S; R,R; or meso; final concentration 250 mM) to radiolabeled DNA in 49 μL TE buffer (10 mM Tris-Cl, 1 mM EDTA, pH 7.1). Incubation was at 37°C for 30 min, followed by ethanol precipitation (22) and lyophilization.

Analysis of Cross-Linking. DEB-treated samples were dissolved in 10 μL of 5 M aqueous urea/0.1% xylene cyanole and loaded onto a 6% denaturing polyacrylamide gel (19:1 acrylamide/bisacrylamide, 42% urea, 30% formamide, 0.35 mm thick, 41 x 37 cm), run on a Hoefer thermojacketed Poker Face gel stand at circa 60 W and ambient temperature. Gels were dried for analysis via phosphorimagery or left wet for purification of alkylated DNA after autoradiography. Percent cross-linking was determined through volume analysis of the family of low-mobility bands in comparison to the single strands with an Amersham Biosciences STORM 840 phosphorimager.

Piperidine Cleavage of Alkylated DNA. Cross-linked (low mobility bands) and monoalkylated DNA (from the higher mobility, single-stranded region of the gels) were recovered from denaturing polyacrylamide gels by the crush-and-soak procedure (22). These samples were cleaved at sites of guanine N7 alkylation by heating at 90°C in 10% aqueous piperidine for 15 min (23). Control (not alkylated) samples showed virtually no degradation under these conditions.

Sequencing Gel Analysis. Samples were dissolved in 10 μL 5 M urea/0.1% xylene cyanole and loaded onto an 8% denaturing gel (19:1 acrylamide/bisacrylamide, 50% urea) run at circa 60 W and 55°C followed by drying and phosphorimagery.

RESULTS

We used a restriction fragment from the 5S RNA gene of Xenopus borealis (Figure 1) to monitor interstrand cross-linking by racemic DEB in comparison to meso, S,S and R,R DEB. This DNA has fifteen potential cross-linking sites (5’-GNC), and its cross-linking by racemic DEB is well characterized (13). Interstrand lesions were analyzed by 6% denaturing polyacrylamide gel electrophoresis (dPAGE). All DEB stereoisomers and the racemic mixture produced the characteristic low-mobility bands diagnostic of interstrand cross-linking (Figure 2). The pattern of low mobility bands was similar for all agents, although the amount of cross-link varied greatly. Following incubation with 250 mM diepoxide, S,S DEB cross-linked about 30% of the total DNA, about twice as much as racemic DEB. R,R and meso DEB were relatively inefficient at producing interstrand cross-links, with meso the least efficient (1.88% cross-linking) of the three isomers.

Figure 1.

Figure 1

Open in a new tab

Sequence of the 3’-end radiolabeled 152 bp fragment used in these studies (20, 21). Potential sites for cross-linking at 5’-GNC sequences are shown in bold. Only one strand is shown for convenience and is numbered from 1 to 152 in the 5’-3’ direction. A single P-32 radiolabel is denoted with an asterisk, although either one or two radiolabels may be incorporated during the fill-in reaction of the EcoRI site.

Figure 2.

Figure 2

Open in a new tab

Interstrand cross-linking patterns of the 152 bp restriction fragment from the 5S RNA gene of Xenopus borealis by S,S DEB (lane 1); R,R DEB (lane 2); meso DEB (lane 3), and racemic DEB (lane 4). Percent cross-linking is as follows: S,S = 28.7%; R,R= 2.17%; meso = 1.88%; racemic = 15.0%.

In order to determine the sites of alkylation for each agent, cross-linked and monoalkylated DNA were extracted from wet denaturing gels and subjected to hot piperidine cleavage. The resulting cleavage products were analyzed via polyacrylamide sequencing gels (Figure 3). For all agents, monoalkylation occurred at every deoxyguanosine residue and cross-linking occurred predominantly at 5’-GNC sites, where N is any base. Traces of cleavage could also be noted at other sites, particularly in the region containing eight consecutive deoxyguanosine residues (G117-G124), suggesting some crosslinking at G-rich sites other than the 5’-GNC consensus sequence. Preferential alkylation at G-rich sites could also lead to monoalkylation of some cross-links, which would contribute to enhanced cleavage in this region. Virtually no high molecular weight material remained, supporting the predominance of cross-linking at piperidine-cleavable sites such as the N7 of guanine.

Figure 3.

Figure 3

Open in a new tab

Piperidine cleavage patterns of monoalkylated (Lanes 1-4) and cross-linked (Lanes 5-8) DNA. Lanes 1 and 6-S,S DEB; Lanes 2 and 7-R,R DEB; Lanes 3 and 8-meso DEB; Lanes 4 and 5-racemic DEB. Landmark deoxyguanosine residues are indicated at the side. Some doubling is visible for low molecular weight fragments because of the possibility of incorporating either one or two deoxyadenosine residues during radiolabeling.

DISCUSSION

Bifunctional electrophiles include such clinically useful anti-tumor drugs as the nitrogen mustards. Although such compounds have many potential cellular targets, anti-cancer potential has been attributed to interstrand cross-linking (24). Indeed, the efficiency of interstrand cross-linking correlates with cytotoxicity for nitrogen mustards (25) and may be a critical aspect governing the action of other antitumor drugs. On the other hand, intrastrand cross-links can be both cytotoxic and mutagenic (16, 26), suggesting that these lesions may contribute to carcinogenic potential.

While S,S DEB is the active form of the prodrug treosulfan used to treat ovarian cancer (6), racemic DEB has been linked to ovarian toxicity and carcinogenicity in mice and rats (27). The dissimilar biological activities of DEB stereoisomers suggest that they may possess distinct reactivity towards DNA. Indeed, S,S DEB has been recently demonstrated to form more 1,3-interstrand cross-links in 5’-GNC trinucleotides than either R,R or meso DEB (18). However, sequence preferences for DNA cross-linking by individual DEB stereoisomers have not previously been compared. Bifunctional carcinogens have unique sequence selectivities. For example, nitrogen mustards link distal guanines within a 5’-GNC context (28, 29) and mitomycin C targets endogenously methylated MeCpG sites (30-32), yet interstrand cross-link formation by N,N’-bis(2-chloroethyl)-nitrosourea is independent of local sequence context (33).

Many previous studies of the sequence preferences for interstrand cross-linking by DEB have used a racemic mixture of stereoisomers (12-14, 34). 5’-GNC was identified as the principal site of DNA cross-linking by racemic DEB, but 5’-GNNC is a secondary target, reacting at about half the frequency of 5’-GNC. In contrast with early predictions (10), the 5’-GC site is poorly cross-linked despite the relatively short length of the butanediol tether. It is likely that significant DNA distortion is required for the formation of 1,3-interstrand 1,4-bis(guan-7-yl)-2,3-butanediol conjugates (34).

In the present study, we explored the role of stereochemistry in the sequence specificity of the interstrand cross-linking reactions of DEB, mapping the sites targeted within a 152 bp restriction fragment previously used for studies with racemic DEB (13). Upon incubation with DEB (S,S; R,R; meso, or racemic), radiolabeled DNA was subjected to dPAGE. Interstrand cross-links appeared as a family of bands with reduced mobility relative to the single strands. Because denaturing gels resolve interstrand cross-linked isomers (35), the pattern of cross-linking can be diagnostic for the sequence preferences of cross-linking (13).

The observed patterns of interstrand cross-links were similar for all types of DEB but the amount of cross-linking varied widely, with the S,S stereoisomer about 15-fold more efficient than either R,R or meso DEB. Racemic DEB, as a mixture of all three isomers, had an intermediate efficiency. These trends are consistent with a recent report of the relative efficiencies of cross-linking with the DEB stereoisomers (18) although our absolute numbers differ. These differences could arise from differences in concentration (100 mM versus 250 mM used in these studies), reaction time (3 hours versus 30 min in these studies), and the target DNA (an oligomer containing a single core 5’-GGC site versus the restriction fragment used in these studies). Subtle differences in flanking sequence effects for the stereoisomers, as noted previously for DEB and diepoxyoctane (14), and secondary sequence preferences could also modulate the relative efficiencies of these agents towards a DNA with many different target sites.

Sites of alkylation were mapped through piperidine cleavage of gel-purified monoadducts and cross-links followed by sequencing gel analysis. The intensities of monoalkylation at different deoxyguanosines were comparable for all types of DEB, with slight variations in intensity between different sites. Preferential alkylation was noted with G-rich regions, such as that from G117-124. Preferential alkylation at runs of contiguous guanines has been noted for other N7 alkylators (36-38), including racemic DEB (13). The sequence preferences for cross-linking also appeared to be comparable for the three optical isomers of DEB, with cross-linking predominating at 5’-GNC sites. Although R,R DEB was recently reported to form interstrand cross-links at 5’-GC sites, these lesions are formed at only about 20% of the efficiency of 5’-GGC cross-links (18). They would therefore be difficult to detect through gel analysis of a DNA with many different target sites such as used in these experiments.

Fairly uniform monoalkylation by racemic DEB has been noted previously (12, 34), indicating that the sequence specificity for cross-linking arises in the second step. That is, after formation of monoadducts, reaction of the second epoxide group occurs preferentially at 5’-GNC sites. We also found similar monoalkylation patterns for each stereoisomer, suggesting that differences in cross-link efficiencies arise from different orientations of reactive groups in stereoisomeric N7-(2’-hydroxy-3’,4’-epoxybut-1’-yl)-guanine (N7-HEBG) intermediates (18). In S,S- and R,R-N7-HEBG, the epoxy oxygen and the 2’-hydroxy group are on the same side of the plane formed by the alkyl chain, while in meso N7-HEBG, the epoxy oxygen and the 2’-hydroxy group are on opposite sides of the plane. It has been proposed that hydrogen bonding between the 2’-hydroxy group in S,S- and R,R-N7-HEBG and the N3 of the 3’-neighboring guanine may position the epoxide ring favorably for the SN2-type nucleophilic attack by the N7 of the guanine in the opposite strand to form an interstrand cross-link (18). Differences in the efficiencies of this reaction for the two stereoisomers may arise from geometry influencing the strengths of these interactions, with the S,S intermediate in better position for the subsequent attack. In contrast, the hydrogen bonding interactions between R,S- and S,R-N7-HEBG intermediates and the 3’-neighboring guanine may better position the epoxide for an SN2 attack by the 3’-guanine to form intrastrand lesions with the meso isomer. Interstrand cross-links also occur at 5’-GNC sites with the meso isomer, although at a reduced efficiency, perhaps due to steric effects. Characterization of the structures of the N7-(2’-hydroxy-3’,4’-epoxybut-1’-yl)-guanine intermediates could confirm the molecular basis for the differences in the lesions formed by the optical isomers of DEB. An understanding of the factors governing lesion formation would be helpful to distinguish the therapeutic and mutagenic effects of DEB.

Because the different optical isomers share the same core preference for interstrand cross-linking, observed differences in their biological activities are more likely to arise from cross-linking efficiencies than sequences targeted. The high efficiency of interstrand cross-linking by S,S DEB is consistent with its potent cytotoxicity and role in the mechanism of treosulfan anti-tumor potential (6). The relatively poor cross-linkers R,R and meso DEB would be expected to have reduced cytotoxicity. However, the intrastrand cross-linking capacity of meso DEB, and to a lesser degree the R,R stereoisomer (18), could contribute to mutagenic effects (39).

ACKNOWLEDGMENT

This work was supported by an NIH Academic Research Enhancement Award from the National Cancer Institute (2R15CA077748-02A1) to JTM, NIH Grant Number P20 RR-016463 from the INBRE Program of the National Center for Research Resources, and a Grant from the National Cancer Institute (CA-100670) to NT.

REFERENCES

  • 1.Ehrenberg L, Hussain S. Genetic toxicity of some important epoxides. Mutation Research. 1981;86:1–113. doi: 10.1016/0165-1110(81)90034-8. [DOI] [PubMed] [Google Scholar]
  • 2.Divine BJ. An update on mortality among workers at a 1,3-butadiene facility. Environ. Health Perspect. 1990;86:119–128. doi: 10.1289/ehp.9086119. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Divine BJ, Wendt JK, Hartman CM. Cancer mortality among workers at a butadiene production facility, in Butadiene and Styrene: Assessment of Health Hazards. In: Sorsa M, Peltonen K, Vainio H, Hemminki K, editors. IARC; Lyon: 1993. pp. 345–362. IARC Scientific Publications No. 127. [PubMed] [Google Scholar]
  • 4.Macaluso M, Larson R, Delzell E, Sathiakumar N, Hovinga M, Julian J, Muir D, Cole P. Leukemia and cumulative exposure to butadiene, styrene, and benzene among workers in the synthetic rubber industry. Toxicology. 1996;113:190–202. doi: 10.1016/0300-483x(96)03444-0. [DOI] [PubMed] [Google Scholar]
  • 5.Albertini R, Clewell H, Himmelstein MW, Morinello E, Olin S, Preston J, Scarano L, Smith MT, Swenberg J, Tice R, Travis C. The use of non-tumor data in cancer risk assessment: reflections on butadiene, vinyl chloride, and benzene. Regulatory Toxicology and Pharmacology. 2003;37:105–132. doi: 10.1016/s0273-2300(02)00019-3. [DOI] [PubMed] [Google Scholar]
  • 6.Hartley JA, O’Hare CC, Baumgart J. DNA alkylation and interstrand cross-linking by treosulfan. Br. J. Cancer. 1999;79:264–266. doi: 10.1038/sj.bjc.6690043. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 7.Cochrane JE, Skopek TR. Mutagenicity of butadiene and its epoxide metabolites: I. Mutagenic potential of 1,2-epoxybutene, 1,2,3,4-diepoxybutane and 3,4-epoxy-1,2-butanediol in cultured human lymphoblasts. Carcinogenesis. 1994;15:713–717. doi: 10.1093/carcin/15.4.713. [DOI] [PubMed] [Google Scholar]
  • 8.Lee DH, Kim T-H, Lee S-Y, Kim H-J, Rhee SK, Yoon B, Pfeifer GP, Lee C-S. Mutations induced by 1,3-butadiene metabolites, butadiene diolepoxide, and 1,2,3,4- diepoxybutane at the Hprt locus in CHO-K1 cells. Mol. Cells. 2002;14:411–419. [PubMed] [Google Scholar]
  • 9.Brookes P, Lawley PD. The alkylation of guanosine and guanylic acid. J. Chem. Soc. 1961:3923. [Google Scholar]
  • 10.Brooks P, Lawley PD. The reaction of mono- and di-functional alkylating agents with nucleic acids. Biochem. J. 1961;80:496–503. doi: 10.1042/bj0800496. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Park S, Tretyakova N. Structural characterization of the major DNA-DNA cross-link of 1,2,3,4-diepoxybutane. Chem. Res. Toxicol. 2004;17:129–136. doi: 10.1021/tx0342058. [DOI] [PubMed] [Google Scholar]
  • 12.Millard JT, White MM. Diepoxybutane cross-links DNA at 5’-GNC sequences. Biochemistry. 1993;32:2120–2124. doi: 10.1021/bi00059a034. [DOI] [PubMed] [Google Scholar]
  • 13.Millard JT, Wilkes EE. Diepoxybutane and diepoxyoctane interstrand cross- linking of the 5S DNA nucleosomal core particle. Biochemistry. 2001;40:10677–10685. doi: 10.1021/bi0109663. [DOI] [PubMed] [Google Scholar]
  • 14.Sawyer GA, Frederick ED, Millard JT. Flanking sequences modulate diepoxide and mustard cross-linking efficiencies at the 5’-GNC site. Chem. Res. Toxicol. 2004;17:1057–1063. doi: 10.1021/tx0499057. [DOI] [PubMed] [Google Scholar]
  • 15.Verly WG, Brakier L, Feit PW. Inactivation of the T7 coliphage by the diepoxybutane stereoisomers. Biochim. Biophys. Acta. 1971;228:400–406. doi: 10.1016/0005-2787(71)90046-3. [DOI] [PubMed] [Google Scholar]
  • 16.Matagne R. Induction of chromosomal aberrations and mutations with isomeric forms of L-threitol-1,4-bismethanesulfonate in plant materials. Mutat. Res. 1969;7:241–247. doi: 10.1016/0027-5107(69)90037-2. [DOI] [PubMed] [Google Scholar]
  • 17.Bianchi A, Contin M. Mutagenic activity of isomeric forms of diepoxybutane in maize. J. Heredity. 1962;53:277–281. [Google Scholar]
  • 18.Park S, Anderson C, Loeber R, Jones R, Tretyakova N. Interstrand and intrastrand DNA-DNA cross-linking by 1,2,3,4-diepoxybutane: role of stereochemistry. J. Am. Chem. Soc. 2005 doi: 10.1021/ja051979x. in press. [DOI] [PubMed] [Google Scholar]
  • 19.Ötvös L, Elekes I. Stereochemistry of the reactions of biopolymers IV. Explanation of stereospecific and asymmetric reactivity of DNA in cross-linking alkylation. Tet. Lett. 1975;29:2481–2484. [Google Scholar]
  • 20.Kuduvalli PN, Townsend CA, Tullius TD. Cleavage by calicheamicin γ I1 of DNA in a nucleosome formed on the 5S RNA gene of Xenopus borealis. Biochemistry. 1995;34:3899–3906. doi: 10.1021/bi00012a005. [DOI] [PubMed] [Google Scholar]
  • 21.Millard JT, Spencer RJ, Hopkins PB. Effect of nucleosome structure on DNA interstrand cross-linking reactions. Biochemistry. 1998;37:5211–5219. doi: 10.1021/bi972862r. [DOI] [PubMed] [Google Scholar]
  • 22.Sambrook J, Fritsch EF, Maniatis T. Molecular Cloning: A Laboratory Manual. Second Edition Cold Spring Harbor Laboratory Press, Cold Spring Harbor; NY: 1989. [Google Scholar]
  • 23.Luce RA, Hopkins PB. Chemical cross-linking of drugs to DNA. Methods Enzym. 2001;340:396–412. doi: 10.1016/s0076-6879(01)40433-2. [DOI] [PubMed] [Google Scholar]
  • 24.Rajski SR, Williams RM. DNA cross-linking agents as antitumor drugs. Chem. Rev. 1998;98:2723–2795. doi: 10.1021/cr9800199. [DOI] [PubMed] [Google Scholar]
  • 25.Sunters A, Springers CJ, Bagshawe KD, Souhami RL, Hartley JA. The cytotoxicity, DNA crosslinking ability and DNA sequence selectivity of the aniline mustards melphalan, chlorambucil, and 4-[bis](2-chloroethyl)amino] benzoic acid. Biochem. Pharm. 1992;44:59–64. doi: 10.1016/0006-2952(92)90038-k. [DOI] [PubMed] [Google Scholar]
  • 26.Yarema KJ, Lippard SJ, Essigmann JM. Mutagenic and genotoxic effects of DNA adducts formed by the anticancer drug cis-diamminedichloroplatinum(II) Nucleic Acids Res. 1995;23:4066–4072. doi: 10.1093/nar/23.20.4066. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 27.Doerr JK, Hollis EA, Sipes IG. Species differences in the ovarian toxicity of 1,3-butadiene epoxides in B6C3F1 mice and Sprague-Dawley rats. Toxicology. 1996;113:128–136. doi: 10.1016/0300-483x(96)03437-3. [DOI] [PubMed] [Google Scholar]
  • 28.Ojwang JO, Grueneberg DA, Loechler EL. Synthesis of a duplex oligonucleotide containing a nitrogen mustard interstrand DNA-DNA cross-link. Cancer Res. 1989;49:6529–6537. [PubMed] [Google Scholar]
  • 29.Millard JT, Raucher S, Hopkins PB. Mechlorethamine cross-links deoxyguanosine residues at 5’-GNC sequences in duplex DNA fragments. J. Am. Chem. Soc. 1990;112:2459–2460. [Google Scholar]
  • 30.Millard JT, Beachy TM. Cytosine methylation enhances mitomycin C cross- linking. Biochemistry. 1993;32:12850–12856. doi: 10.1021/bi00210a038. [DOI] [PubMed] [Google Scholar]
  • 31.Johnson WS, He Q-Y, Tomasz M. Selective recognition of the m5 CpG dinucleotide sequence in DNA by mitomycin C for alkylation and cross-linking. Bioorg. Med. Chem. 1995;3:851–860. doi: 10.1016/0968-0896(95)00067-q. [DOI] [PubMed] [Google Scholar]
  • 32.Li V-S, Reed M, Zheng Y, Kohn H, Tang M. C5 cytosine methylation at CpG sites enhances sequence selectivity of mitomycin C-DNA bonding. Biochemistry. 2000;39:2612–2618. doi: 10.1021/bi991307h. [DOI] [PubMed] [Google Scholar]
  • 33.Fischhaber PL, Gall AS, Duncan JA, Hopkins PB. Direct demonstration in synthetic oligonucleotides that N,N’-bis(2-chloroethyl)-nitrosourea cross-links N1 of deoxyguanosine to N3 of deoxycytidine on opposite strands of duplex DNA. Cancer Res. 1999;59:4363–4368. [PubMed] [Google Scholar]
  • 34.Yunes MJ, Charnecki SE, Marden JJ, Millard JT. 1,2,5,6-Diepoxyhexane and 1,2,7,8-diepoxyoctane cross-link duplex DNA at 5’-GNC sequences. Chem. Res. Toxicol. 1996;9:994–1000. doi: 10.1021/tx960059c. [DOI] [PubMed] [Google Scholar]
  • 35.Millard JT, Weidner MF, Kirchner JJ, Ribeiro S, Hopkins PB. Sequence preferences of DNA interstrand crosslinking agents: quantitation of interstrand crosslink locations in DNA duplex fragments containing multiple crosslinkable sites. Nucleic Acids Res. 1991;19:1885–1891. doi: 10.1093/nar/19.8.1885. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36.Mattes WB, Hartley JA, Kohn KW. DNA sequence selectivity of guanine-N7 alkylation by nitrogen mustards. Nucleic Acids Res. 1986;14:2971–2987. doi: 10.1093/nar/14.7.2971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 37.Hartley JA, Gibson NW, Kohn KW, Mattes WB. DNA sequence selectivity of guanine-N7 alkylation by three antitumor chloroethylating agents. Cancer Res. 1986;46:1943–1947. [PubMed] [Google Scholar]
  • 38.Said B, Shank RC. Nearest neighbor effects on carcinogen binding to guanine runs in DNA. Nucleic Acids Res. 1991;19:1311–1316. doi: 10.1093/nar/19.6.1311. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 39.Carmical JR, Kowalczyk A, Zou Y, Van Houten B, Nechev LV, Harris CM, Harris TM, Lloyd RS. Butadiene-induced intrastrand DNA cross-links: a possible role in deletion mutagenesis. J. Biol. Chem. 2000;275:19482–19489. doi: 10.1074/jbc.M002037200. [DOI] [PubMed] [Google Scholar]

RESOURCES