Mechanism of stimulation of the DNA glycosylase activity of hOGG1 by the major human AP endonuclease: bypass of the AP lyase activity step

The DNA glycosylase NEIL3 has been implicated in DNA repair pathways including the base excision repair and the interstrand cross-link repair pathways via its DNA glycosylase and/or AP lyase activity, which are considered canonical roles of NEIL3 in genome integrity. Compared with the other DNA glycosylases NEIL1 and NEIL2, Xenopus laevis NEIL3 C terminus has two highly conserved zinc finger motifs containing GRXF residues (designated as Zf-GRF). It has been demonstrated that the minor AP endonuclease APE2 contains only one Zf-GRF motif mediating interaction with single-strand DNA (ssDNA), whereas the major AP endonuclease APE1 does not. It appears that the two NEIL3 Zf-GRF motifs (designated as Zf-GRF repeat) are dispensable for its DNA glycosylase and AP lyase activity; however, the potential function of the NEIL3 Zf-GRF repeat in genome integrity remains unknown. Here, we demonstrate evidence that the NEIL3 Zf-GRF repeat was associated with a higher affinity for shorter ssDNA than one single Zf-GRF motif. Notably, our protein–protein interaction assays show that the NEIL3 Zf-GRF repeat but not one Zf-GRF motif interacted with APE1 but not APE2. We further reveal that APE1 endonuclease activity on ssDNA but not on dsDNA is compromised by a NEIL3 Zf-GRF repeat, whereas one Zf-GRF motif within NEIL3 is not sufficient to prevent such activity of APE1. In addition, COMET assays show that excess NEIL3 Zf-GRF repeat reduces DNA damage in oxidative stress in Xenopus egg extracts. Together, our results suggest a noncanonical role of NEIL3 in genome integrity via its distinct Zf-GRF repeat in suppressing APE1 endonuclease-mediated ssDNA breakage.

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Mutant E.Coli 3-methyladenine DNA glycosylase II reveals AP-lyase activity

Seibutsu Butsuri ◽

10.2142/biophys.40.s29_1 ◽

2000 ◽

Vol 40 (supplement) ◽

pp. S29

Author(s):

H. Nakano ◽

H. Matsuda ◽

Y. Yamagata ◽

M. Sekiguchi ◽

Y. Kobayashi

Keyword(s):

Dna Glycosylase ◽

Lyase Activity ◽

Ap Lyase

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The use of thioglycollate to demonstrate DNA AP (apurinic/apyrimidinic-site) lyase activities. Biological consequences of thiol addition to the 5′ product of a β-elimination reaction at an AP site in DNA

Biochemical Journal ◽

10.1042/bj2730777 ◽

1991 ◽

Vol 273 (3) ◽

pp. 777-782 ◽

Cited By ~ 2

Author(s):

S Bricteux-Grégoire ◽

W G Verly

Keyword(s):

Escherichia Coli ◽

Elimination Reaction ◽

Ap Endonuclease ◽

Lyase Activity ◽

Ap Sites ◽

Ap Lyase ◽

Ap Site

Thioglycollate reacts with the 5′ product of AP lyase activity on apurinic/apyrimidinic (AP) sites in DNA. The 3′-terminal thioglycollate-unsaturated sugar 5-phosphate adduct can be released by the use of Escherichia coli endonuclease IV or endonuclease VI, and identified by DEAE-Sephadex chromatography. In contrast, the mammalian AP endonuclease is unable to excise a 3′-terminal thiol-unsaturated sugar adduct; this lesion, which must sometimes occur in vivo, might be irreparable and have pathological consequences.

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Nonenzymatic release of N7-methylguanine channels repair of abasic sites into an AP endonuclease-independent pathway in Arabidopsis

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1719497115 ◽

2018 ◽

Vol 115 (5) ◽

pp. E916-E924 ◽

Cited By ~ 4

Author(s):

Casimiro Barbado ◽

Dolores Córdoba-Cañero ◽

Rafael R. Ariza ◽

Teresa Roldán-Arjona

Keyword(s):

Excision Repair ◽

Biochemical Properties ◽

Abasic Site ◽

Single Nucleotide ◽

Ap Endonuclease ◽

Abasic Sites ◽

Lyase Activity ◽

Ap Sites ◽

Base Excision ◽

Ap Lyase

Abasic (apurinic/apyrimidinic, AP) sites in DNA arise from spontaneous base loss or by enzymatic removal during base excision repair. It is commonly accepted that both classes of AP site have analogous biochemical properties and are equivalent substrates for AP endonucleases and AP lyases, although the relative roles of these two types of enzymes are not well understood. We provide here genetic and biochemical evidence that, in Arabidopsis, AP sites generated by spontaneous loss of N7-methylguanine (N7-meG) are exclusively repaired through an AP endonuclease-independent pathway initiated by FPG, a bifunctional DNA glycosylase with AP lyase activity. Abasic site incision catalyzed by FPG generates a single-nucleotide gap with a 3′-phosphate terminus that is processed by the DNA 3′-phosphatase ZDP before repair is completed. We further show that the major AP endonuclease in Arabidopsis (ARP) incises AP sites generated by enzymatic N7-meG excision but, unexpectedly, not those resulting from spontaneous N7-meG loss. These findings, which reveal previously undetected differences between products of enzymatic and nonenzymatic base release, may shed light on the evolution and biological roles of AP endonucleases and AP lyases.

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