5,6-Dihydrothymid-5-yl (4) is generated via Norrish type I cleavage of isopropyl ketone 7. Ketone 7 was site specifically incorporated into chemically synthesized polythymidylates and an oligonucleotide containing all four native deoxyribonucleotides. No damage is induced in oligonucleotides containing 7 upon photolysis under anaerobic conditions. In the presence of O2, strand breaks and alkaline labile lesions are formed at the original site of 7, and at nucleotides adjacent to the 5‘-phosphate of 7. Kinetic isotope effect experiments reveal that direct strand scission at the thymidine adjacent to the 5‘-phosphate of 4 arises from C1‘ hydrogen atom abstraction. The observed KIE (∼3.9) is attributed to hydrogen atom abstraction from C1‘ by the peroxyl radical 35 derived from 4. Enzymatic end group analysis and measurement of free base release are consistent with a process involving C1‘ hydrogen atom abstraction. Cleavage experiments carried out in the presence of t-BuOH (1.05 M) and NaN3 (10 mM) indicate that damage does not result from hydroxyl radical, but that 1O2 is responsible for a significant amount of the observed strand damage.

A contrathermodynamic sequence selectivity (5′-deoxyadenosine > 5′-deoxyguanosine) for UVirradiation-induced strand damage in duplex DNA containing 5-bromo-2′-deoxyuridine was reported several years ago (Saito, I.; Sugiyama, H. J. Am. Chem. Soc. 1990, 112, 6720.). In contrast, much smaller sequence selectivity was observed for similar duplexes containing 5-iodo-2′-deoxyuridine. We investigated the mechanism of UV-irradiation-induced cleavage of duplex DNA containing 5-bromo-2′-deoxyuridine (1, BrdU) and 5-iodo-2′-deoxyuridine (2, IdU) under anaerobic conditions using a variety of structural probes. The preference for UV-induced cleavage in 5′-dABrdU sequences is a confluence of at least three factors, photoinduced forward electron transfer, charge recombination, and electron migration within the DNA duplex. Our results also indicate that UV-irradiation of duplexes (32 nucleotides long) containing 5-iodo-2′-deoxyuridine results in strand scission involving initial photoinduced single electron transfer. The selectivity for 5′-dAIdU sequences is smaller than that in the analogous 5-bromo-2′-deoxyuridine duplexes and may be the result of faster dehalogenation of the initially formed 5-halopyrimidine radical anion and/or competitive direct carbon-iodine bond homolysis.5-Bromo-2′-deoxyuridine (1, BrdU) and 5-iodo-2′-deoxyuridine (2, IdU) exhibit a number of interesting and potentially useful chemical properties. 1 For instance, incorporation of these molecules in nucleic acids sensitizes the biopolymers to γ-radiolysis. 2 In addition, the 5-halopyrimidine nucleosides' sensitivity to UV-irradiation has been exploited in the application of these molecules as structural probes of protein-nucleic acid interactions and nucleic acid structure. 3,4 The enhancement of DNA damage caused by UV-irradiation of duplexes containing 1 and 2 has received considerable attention. 5-8 Approximately 10 years ago Saito and Sugiyama reported that UV-induced strand damage in duplex DNA containing 1 was highly dependent on the identity of the nucleotide bonded to the 5′-phosphate of 5-bromo-2′-deoxyuridine. 5 In contrast to previous studies, it was observed that UV-irradiated duplexes containing the sequence 5′-dABrdU exhibited significantly greater amounts of alkali-labile lesion formation than analogous molecules containing either a 2′-deoxyguanosine (dG) or 2′-deoxycytidine (dC) in place of the adjacent 2′-deoxyadenosine (dA). 9 These researchers postulated a novel mechanism involving initial photoinduced single electron transfer (PSET) from a 2′-deoxyadenosine bonded to the 5′-phosphate of 1 (Scheme 1). Preferential damage observed in 5′-dABrdU (3) sequences via PSET compared to those containing a 2′-deoxyguanosine nucleotide bonded to the 5′-phosphate of the halopyrimidine was surprising, given that electron transfer from deoxyguanosine is more favorable thermodynamically. 10 We now wish to report on studies that support this proposal, and demonstrate that the origin of this unexpected sequence selectivity for DNA damage in biopolymers ...

The response of mutagenicity to the stepwise replacement of chlorine atoms and the hydroxyl group by hydrogen in 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX, 1) was determined in several assays by using Salmonella typhimurium tester strain (TA100). In all, eight MX derivatives were assayed. Several were studied together in at least one assay. In addition to MX, the seven included 3-chloro-4-(dichloromethyl)-2(5H)-furanone (RMX, 2), 3-chloro-4-(chloromethyl)-5-hydroxy-2(5H)-furanone (3), 3-chloro-4-(chloromethyl)-2(5H)-furanone (4), 4-(chloromethyl)-5-hydroxy-2(5H)-furanone (5), 4-chloromethyl-2(5H)-furanone (6), and 4-(dichloromethyl)-2(5H)-furanone (8). Compounds 1-6 were mutagenic. Compound 8 gave erratic results. 4-(Acetoxymethyl)-2(5H)-furanone (11) was nonmutagenic. The largest drop in mutagenicity amounted to a factor of about 10(2) for the replacement of the hydroxyl group or a C-3 chlorine atom from 3. Other replacements of the hydroxyl group or a C-3 or C-6 chlorine atom amounted to mutagenicity diminished by a factor of only 10. On the basis of the rates of UV spectral changes under assay conditions, chemical half-life values (ct 1/2) for 1-6 and 8 were estimated as indicators of compound stability. However, mutagenicity differences were shown to result principally from the intrinsic mutagenicities of the six compounds 1-6 rather than from differences in stability.(ABSTRACT TRUNCATED AT 250 WORDS)

The sodium borohydride reduction of 3,4-dichloro-5-hydroxy-2(5H)-furanone (mucochloric acid) resulted in the formation of 3,4-dichloro-2(5H)-furanone and 4-chloro-2(5H)-furanone formed in relative amounts of 98 and 2%, respectively. Mucochloric acid and 3,4-dichloro-2(5H)furanone were assayed together against Salmonella fyphimurium (TA100). Mucochloric acid was consistently more mutagenic than 3,4-dichloro-2(5H)-furanone by a factor ranging from 36 to 49 times as established from three determinations of relative mutagenicity. Thus, the hydroxyl group substituted at the 5 position is concluded to have a marked influence on the mutagenicity of a chlorinated 2(5H)-furanone.

The mutagenicities of 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX, compound 1), 3-chloro-4-(dichloromethyl)-2(5H)-furanone (RMX, compound 6), and 2-(dichloromethyl)-3,3-dichloropropenal (TCB, compound 7) were determined in the same assay and in repetitive determinations using Salmonella typhimurium (TA 100) without microsomal fraction activation. In addition, the mutagenicity of 2-methyl-3,3-dichloropropenal (compound 8) was assayed in the same manner although not simultaneously with MX, RMX, and TCB. This study was undertaken to ascertain the role of open- and closed-ring forms of MX in the mutagenicity of MX. MX proved to be roughly 100 times more mutagenic than the open-ring analogue TCB and 10 times more mutagenic than the closed-ring analogue RMX. Compound 8 was inactive. Assay stability of the three active compounds in Vogel-Bonner medium at 38 degrees C was estimated as the chemical half-life values by following the change in UV absorbance at selected wave lengths. Half-life values were 10.7, 2.6, and 2.8 hr, respectively, for MX, RMX, and TCB. The enhanced mutagenicity of MX relative to RMX and TCB is attributed to the intrinsic mutagenicity of MX and its greater stability is judged to play only a minor role. Moreover, the greater mutagenicity of the closed-ring analogue RMX relative to the open-ring analogue TCB points to the ring form of MX as the active species even though the open form of MX is predominant under assay conditions.