Sequence specificity of DNA-psoralen photoproduct formation in supercoiled plasmid DNA (pUC19)

Abstract

Supercoiled pUC19 DNA, photoreacted with psoralen derivatives (xanthotoxin (8-methoxypsoralen, 8-MOP), 4,5′,8-trimethylpsoralen (TMP) and angelicin), influences the enzymatic activity of restriction enzymes in a different manner, although all the enzymes employed contain, within their recognition sites, suitable nucleic acid bases for photoproduct formation. The activity of the enzymes is strongly influenced by the photomodification of thymine residues within their recognition sites. 5′-TpA sequences favour intercalation as an essential prerequisite of the photoreaction, while 5′ApT sequences do not. This, in turn, influences photoproduct formation and the inhibition of the action of the restriction enzymes KpnI, SspI, DraI and RsaI, but not EcoRI and BamHI. The inhibitory effect is independent of the number of cleavage sites and also of whether monoaddition products or crosslinks are formed. Psoralen intercalation alone does not affect the activity of the restriction enzymes used.