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Purification and characterization of a novel deoxyinosine-specific enzyme, deoxyinosine 3' endonuclease, from Escherichia coli

Purification and characterization of a novel deoxyinosine-specific enzyme, deoxyinosine 3' endonuclease, from Escherichia coli

Journal of Biological Chemistry 269(23): 16260-8

We have purified a novel endonuclease from Escherichia coli that recognizes deoxyinosine, a deamination product of deoxyadenosine in DNA. This activity, which we named deoxyinosine 3' endonuclease, is different from the known hypoxanthine DNA N-glycosylases which have been partially characterized in E. coli and other organisms. The enzyme was purified 24,800-fold to apparent homogeneity. SDS- and activity PAGE analyses indicate that the enzyme has an apparent molecular mass of 25 kDa. Deoxyinosine 3' endonuclease recognized deoxyinosine in both single- and double-stranded DNA but exhibited a 4-fold preference for double stranded over single-stranded DNA. In addition to deoxyinosine, the enzyme recognized urea residues and A-P sites. Deoxyinosine 3' endonuclease has an obligatory requirement for Mg-2+, but other cations such as Co-2+ and Mn-2+ could partially replace Mg-2+. The optimal pH for deoxyinosine 3' endonuclease was around 7.5. In contrast to most of the known repair enzymes, deoxyinosine 3' endonuclease makes an incision at the second phosphodiester bond 3' to a deoxyinosine or A-P site, leaving behind the intact lesion on the nicked DNA- Therefore, deoxyinosine 3' endonuclease recognizes, but does not remove, the lesion from the DNA molecule. The biological significance of this novel activity is discussed with reference to other repair activities in E. coli.

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Accession: 009276564

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PMID: 8206931

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