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Mammalian nucleotide excision repair (NER) eliminates carcinogen-DNA adducts by double endonucleolytic cleavage and subsequent release of 24-32 nucleotide-long single-stranded fragments. Here we manipulated the deoxyribose-phosphate backbone of DNA to analyze the mechanism by which damaged strands are discriminated as substrates for dual incision. We found(More)
A general scheme is described for the in vitro evolution of protein catalysts in a biologically amplifiable system. Substrate is covalently and site specifically attached by a flexible tether to the pIII coat protein of a filamentous phage that also displays the catalyst. Intramolecular conversion of substrate to product provides a basis for selecting(More)
BACKGROUND Deoxyribosylnucleotide radicals with a radical center at the 4'-position are important intermediates in radical-induced DNA strand cleavage. In the presence of O2, these DNA radicals yield cleavage products that are partly oxidized. In the past, the postulated peroxide intermediates could not be detected directly because they were unstable under(More)
Many genotoxic agents form base lesions that inhibit DNA polymerases. To study the mechanism underlying termination of DNA synthesis on defective templates, we tested the capacity of a model enzyme (Klenow fragment of Escherichia coli DNA polymerase I) to catalyze primer elongation across a series of C4' deoxyribose derivatives. A site with inverted C4'(More)
BACKGROUND To gain a molecular understanding of a biochemical process, the crystal structure of enzymes that catalyze the reactions involved is extremely helpful. Often the question arises whether conformations obtained in this way appropriately reflect the reactivity of enzymes, however. Rates that characterize transitions are therefore compulsory(More)
BACKGROUND The genetic integrity of living organisms is maintained by a complex network of DNA repair pathways. Nucleotide excision repair (NER) is a versatile process that excises bulky base modifications from DNA. To study the substrate range of this system, we constructed bulky deoxyribose adducts that do not affect the chemistry of the corresponding(More)
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