Psoralen--lexitropsin hybrids: DNA sequence selectivity of photoinduced cross-linking from MPE footprinting and exonuclease III stop assay, and mode of binding from electric linear dichroism.
The properties of certain hybrids 3 and 5 bearing a photoactivatable psoralen group attached to DNA sequence recognizing lexitropsin carriers have been examined. The hybrids bind to poly(dA-dT) with Kapp of 2.8 and 0.9 x 10(7) M-1, i.e. greater than or equal to that of netropsin (Kapp = 1.0 x 10(7) M-1), indicating that the psoralen moiety may contribute to binding in the case of 5. Photoinduced cross-linking of DNA by 3 and 5, while efficient, is less so than that of individual psoralens and reaches a maximum at a ligand to DNA base pair ratio (r) of 0.2. Complementary strand methidium-propyl-EDTA (MPE).Fe(II) footprinting demonstrated that, in the dark, the sequence preferential recognition of hybrids 3 and 5 is dominated by the lexitropsin moiety. Examination of 360 nm photoinduced DNA cross-linking by the hybrids 3 and 5 was carried out using an exonuclease III stop assay. This revealed that > 95% of the DNA remained double stranded, indicating that 3 and 5 generate primarily biadducts at AT-rich sequences. This assay also located individual monoadduct sites, some of which are remote from the dominant cross-linked sites. When the samples were exposed to 254 nm UV light before loading onto the gel to reverse the photoproducts, the pattern of the exonuclease III stop bands was not altered significantly compared with the experiment without 254 nm irradiation. It is concluded that these termination sites include both mono- and biadducts. Electric linear dichroism examination of the DNA complexes of hybrids 3 and 5 (without light activation) provides evidence that the lexitropsin portion binds in the minor groove, while the psoralen portion intercalates in a suitably located site for subsequent photoinduced cross-linking.
Université de Montpellier II, Sciences et Techniques du Languedoc, Laboratoire de Chimie Bio-Organique, URA CNRS, France.
K E
KE
Rao
G
G
Gosselin
D
D
Mrani
C
C
Périgaud
J L
JL
Imbach
C
C
Bailly
J P
JP
Hénichart
P
P
Colson
C
C
Houssier
J W
JW
Lown
D000970:Antineoplastic Agents / Q000378:metabolism*; D001483:Base Sequence; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004492:Edetic Acid / Q000031:analogs & derivatives* / Q000378:metabolism; D005090:Exodeoxyribonucleases / Q000494:pharmacology; D011564:Furocoumarins / Q000378:metabolism*; D008969:Molecular Sequence Data; D009429:Netropsin / Q000031:analogs & derivatives* / Q000378:metabolism
D000970:Antineoplastic Agents; D011564:Furocoumarins; C038200:methidiumpropyl-EDTA-iron(II); C053654:lexitropsin; D009429:Netropsin; D004247:DNA; D004492:Edetic Acid; D005090:Exodeoxyribonucleases; C020133:exodeoxyribonuclease III