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THE SITE‐SPECIFIC INHIBITION OF Bgl I CLEAVAGE BY PSORALEN PHOTOADDUCTS
Author(s) -
Ostrander Elaine A.,
Robinson Gordon W.,
Isaacs Stephen T.,
Tessman John,
Hallick Lesley M.
Publication year - 1986
Publication title -
photochemistry and photobiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.818
H-Index - 131
eISSN - 1751-1097
pISSN - 0031-8655
DOI - 10.1111/j.1751-1097.1986.tb03559.x
Subject(s) - cleavage (geology) , psoralen , furocoumarins , dna , chemistry , pbr322 , recognition sequence , stereochemistry , base pair , adduct , restriction enzyme , biochemistry , biology , photochemistry , plasmid , paleontology , organic chemistry , fracture (geology)
— We have investigated the site specificity of furocoumarins by using fluorescent densitometry to examine the frequency of cleavage by the restriction enzyme Bgl I. This enzyme has an 11 base pair (bp) recognition sequence which varies slightly from site to site because it includes a 5 base pair neutral region. Cleavage at all three Bgl I recognition sites in pBR322 was inhibited by the photoaddition of the psoralen derivative 4′‐hydroxymethyl‐4,5′,8‐trimethylpsoralen (HMT) which forms both crosslinks and monoad‐ducts in a dose‐dependent manner. One site, which contains two thymidines in a crosslinkable configuration, was observed to be markedly more sensitive to HMT photoadducts. In contrast Bgl I cleavage at all sites was relatively resistant to the derivative 5‐methylisopsoralen (5‐MIP), which forms only monoadducts. When HMT‐reacted DNA was generated with widely different ratios of monoad‐ducts to crosslinks (3% and 40% crosslinks), essentially the same level and pattern of inhibition was observed in both cases. Taken together, the data imply that differences in inhibition seen at the three cutting sites of Bgl I with HMT are attributable to DNA sequence and the role it plays in adduct positioning.