Journal Article

Rates of base excision repair are not solely dependent on levels of initiating enzymes

Enrico Cappelli, Tapas Hazra, Jeff W. Hill, Geir Slupphaug, Massimo Bogliolo and Guido Frosina

in Carcinogenesis

Volume 22, issue 3, pages 387-393
Published in print March 2001 | ISSN: 0143-3334
Published online March 2001 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/22.3.387
Rates of base excision repair are not solely dependent on levels of initiating enzymes

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The oxidized base 8-oxo-7,8-dihydroguanine (8-oxoG), the product of deamination of cytosine uracil (U), and the sites of base loss [abasic (AP) sites] are among the most frequent mutagenic lesions formed in the human genome under physiological conditions. In human cells, the enzymatic activities initiating DNA base excision repair (BER) of 8-oxoG, U and AP sites are the 8-oxoG DNA glycosylase (hOGG1), the U-DNA glycosylase (UNG) and the major hydrolytic AP endonuclease (APE/HAP1), respectively. In recent work, we observed that BER of the three lesions occurs in human cell extracts with different efficacy. In particular, 8-oxoG is repaired on average 4-fold less efficiently than U, which, in turn, is repaired 7-fold slower than the natural AP site. To discriminate whether the different rates of repair may be linked to different expression of the initiating enzymes, we have determined the amount of hOGG1, UNG and APE/HAP1 in normal human cell extracts by immunodetection techniques. Our results show that a single human fibroblast contains 123 000 ± 22 000 hOGG1 molecules, 178 000 ± 20 000 UNG molecules and 297 000 ± 50 000 APE/HAP1 molecules. These limited differences in enzyme expression levels cannot readily explain the different rates at which the three lesions are repaired in vitro. Addition to reaction mixtures of titrated amounts of purified hOGG1, UNG and APE/HAP1 variably stimulated the in vitro repair replication of 8-oxoG, U and the AP site respectively and the increase was not always proportional to the amount of added enzyme. We conclude that the rates of BER depend only in part on cellular levels of initiating enzymes.

Keywords: AP, abasic; APE/HAP1, human AP endonuclease; AU, absorbance units; BER, base excision repair; hOGG1, 8-oxoG DNA glycosylase; 8-oxoG, 8-oxo-7,8-dihydroguanine; U, uracil; UNG, U-DNA glycosylase; UNGΔ84, human recombinant UNG; UNG1, mitochondrial form of human UNG; UNG2, nuclear form of human UNG.

Journal Article.  5390 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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