Two distinct human DNA diesterases that hydrolyze 3′-blocking deoxyribose fragments from oxidized DNA

Abstract

Mammalian cells were investigated for enzymes that help correct oxidative damages in DNA. We focused on 3'-repair diesterases, which process DNA ends at oxidative strand breaks by removing 3'-blocking fragments of deoxyribose that prevent DNA repair synthesis. Two enzymes were found in a variety of mouse, bovine and human tissues and cultured cells. The two activities were purified to differing degrees from HeLa cells. One enzyme had the properties of the known HeLa AP endonuclease (Mr approximately 38,000, with identical substrate specificity and reaction requirements, and cross-reactivity with anti-HeLa AP endonuclease antiserum) and is presumed identical to that protein. The second activity did not interact with anti-HeLa AP endonuclease antibodies and had relatively less AP endonuclease activity. This second enzyme may have been detected in other studies but never characterized. In addition to the 3'-repair diesterase and AP endonuclease, this partially purified preparation also harbored DNA 3'-phosphatase and 3'-deoxyribose diesterase activities. It is unknown whether all activities detected in the second preparation are due to a single protein, although activity against undamaged DNA was not detected. The in vivo roles of these two widely distributed 3'-repair diesterase/AP endonucleases have not been determined, but with the characterizations presented here such questions may now be focused.

Chimeric adenoviral vector Ad5/F35-mediated APE1 siRNA enhances sensitivity of human colorectal cancer cells to radiotherapy in vitro and in vivo

“…APE1 is the major AP endonuclease in human cells and accounts for 95% of the total AP endonuclease activity, and is essential for the protection of cells against the toxic effects of several classes of DNAdamaging agents. [7][8][9][10] APE1 is active in both and interacts with several proteins including 8-oxoguanine DNA glycosylase , X-ray cross-complementing-1 , proliferating cell nuclear antigen, flap endonuclease 1 and polymerase b. 32 In addition to AP endonuclease activity, APE1 also possesses 3 0 -phosphodiesterase, 3 0 -phosphatase and 3 0 -5 0 -exonuclease functions specific for the 3 0 termini of internal nicks and gaps in DNA.…”

“…[4][5][6] The human apurinic/apyrimidinic endonuclease (APE1) is an essential enzyme in the base excision repair (BER) pathway, which is responsible for the repair of oxidative and alkylation DNA damage. APE1 accounts for nearly all of the abasic site cleavage activity in most cultured human cell lines, [7][8][9][10] and is essential for the protection of cells against the toxic effects of several classes of DNA-damaging agents. Moreover, APE1 has 3 0 -repair diesterase or phosphatase activity, albeit at 200-fold lower than its AP endonuclease activity.…”

Human Apurinic/Apyrimidinic Endonuclease 1

“…The initial work on the purified mouse, bovine, and human AP endonuclease found that the mammalian protein exhibited not only a powerful class II AP site incision activity, but also the ability to excise 3¢-damages, such as 3¢-phosphates, 3¢-phosphoglycolate esters, and 3¢-deoxyribose fragments (27,173,182,183). These 3¢-end blocking groups prevent primer extension by a DNA polymerase or nick ligation by a DNA ligase, and, thus, have the potential to promote chromosome instability as unrepaired strand breaks.…”

Dynamics and diversions in base excision DNA repair of oxidized abasic lesions

“…In view of the amount of Ape1 present in most cell types (Cappelli et al, 2001;Chen et al, 1991), it appeared likely that most dL residues would be 5' cleaved by the enzyme before AP lyases would have the opportunity to react. The next enzyme of BER, Polb, would then be poised to mediate excision of the abasic residue.…”