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I. Ponten ; Seog K. Kim (Institutionen för fysikalisk kemi) ; A. Graslund ; Bengt Nordén (Institutionen för fysikalisk kemi) ; B. Jernstrom
Carcinogenesis (0143-3334). Vol. 15 (1994), 10, p. 2207-2213.
[Artikel, refereegranskad vetenskaplig]

The oligonucleotide 5'-d(CCTATAGATATCC) has been reacted with the (+)- or (-)-enantiomers of trans-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(+)-and (-)-anti-BPDE respectively]. Consistent with previous studies employing single-stranded oligonucleotides, adduct formation of both anti-BPDE enantiomers preferentially involved trans-addition of the C10 position of the diol-epoxide to the exocyclic nitrogen of deoxyguanosine [in the following abbreviated as (+)-BPDE(t)-N-2-G and (-)-BPDE(t)-N-2-G adducts respectively]. The unmodified or (+)-BPDE(t)-N-2-G-modified oligonucleotide was allowed to form duplexes with the complementary sequence 5'-d(GGATATCTATAGG) or sequences in which C has been replaced with T, G or A and analysed with regard to thermal stability. The presence of a (+)-BPDE(t)-N-2-G adduct in oligonucleotide duplexes substantially decreased the value of the melting point relative to the corresponding unmodified duplex. In mismatched complexes containing the (+)-BPDE(t)-N-2-G adduct, a further decrease in thermal stability was observed. The presence of a (+)-BPDE(t)-N-2-G adduct did not seem to change the extent of hyperchromicity (approximate to 20%) upon melting. 5'-d(GGATATCTATAGG) or strands in which C was replaced with T,G or A were gradually added to (+)- or (-)-BPDE(t)-N-2-G-modified oligonucleotides and the fluorescence emission intensity was determined. In all cases with (+)-BPDE(t)-N-2-G, except when C was replaced with A in the complement, the fluorescence intensity steadily decreased and became constant at equal strand concentrations. When a strand containing A in place of C was gradually added to the (+)-BPDE(t)-N-2-G oligonucleotide, a marked increase in the fluorescence intensity was observed (>3-fold). In contrast, addition of strands containing A, T or G to the (-)-BPDE(t)-N-2-G-modified oligonucleotide increased tbe fluorescence intensity from 1.5- to >5-fold. Addition of the fully complementary sequence to the (-)-BPDE(t)-N-2-G-containing oligonucleotide resulted in reduced fluorescence, however less pronounced than with the (+)-BPDE(t)-N-2-G-modified analogue. Significant changes in spectral properties of the adducts were observed in the duplexes. The absorption and fluorescence excitation maxima of the single-stranded (+)-BPDE(t)-N-2-G-modified oligonucleotide were at 353 nm. Insertion of C or A opposite the adduct caused a significant shift of these maxima to shorter wavelengths (347-348 nm). Addition of acrylamide, a fluorescence quencher, reduced the fluorescence intensity in all cases, but to variable extents. The adducts not quenchable by acrylamide demonstrate spectral properties similar to those of the single-stranded (+)-BPDE(t)-N-2-G-modified oligonucleotide.

Denna post skapades 2011-08-10.
CPL Pubid: 144019


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