A Single Engineered Point Mutation in the Adenine Glycosylase MutY Confers Bifunctional Glycosylase/AP Lyase Activity†

Biochemistry ◽  
2000 ◽  
Vol 39 (33) ◽  
pp. 10098-10109 ◽  
Author(s):  
Scott D. Williams ◽  
Sheila S. David
Keyword(s):  
Point Mutation ◽  
Lyase Activity ◽  
Ap Lyase ◽  
Adenine Glycosylase

Adenine Glycosylase MutY of Corynebacterium pseudotuberculosis presents the antimutator phenotype and evidences of glycosylase/AP lyase activity in vitro

2016 ◽  
Vol 44 ◽  
pp. 318-329 ◽  
Author(s):  
Rafael Cançado de Faria ◽  
Liliane Gonçalves Vila-Nova ◽  
Mainá Bitar ◽  
Bruno Carvalho Resende ◽  
Larissa Sousa Arantes ◽  
...  
Keyword(s):  
Corynebacterium Pseudotuberculosis ◽  
Lyase Activity ◽  
Ap Lyase ◽  
Adenine Glycosylase

scholarly journals A non-canonical role for the DNA glycosylase NEIL3 in suppressing APE1 endonuclease-mediated ssDNA damage

2020 ◽  
Vol 295 (41) ◽  
pp. 14222-14235 ◽  
Author(s):  
Anh Ha ◽  
Yunfeng Lin ◽  
Shan Yan
Keyword(s):  
Excision Repair ◽  
Dna Glycosylase ◽  
Genome Integrity ◽  
Ap Endonuclease ◽  
C Terminus ◽  
Egg Extracts ◽  
Lyase Activity ◽  
Repair Pathways ◽  
Base Excision ◽  
Ap Lyase

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.

scholarly journals Human Nei-like protein NEIL3 has AP lyase activity specific for single-stranded DNA and confers oxidative stress resistance inEscherichia colimutant

2009 ◽  
Vol 14 (2) ◽  
pp. 261-270 ◽  
Author(s):  
Masashi Takao ◽  
Yoshitsugu Oohata ◽  
Kengo Kitadokoro ◽  
Kumiko Kobayashi ◽  
Shigenori Iwai ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Stress Resistance ◽  
Single Stranded Dna ◽  
Oxidative Stress Resistance ◽  
Lyase Activity ◽  
Ap Lyase

scholarly journals Requirements for 5′dRP/AP lyase activity in Ku

2014 ◽  
Vol 42 (17) ◽  
pp. 11136-11143 ◽  
Author(s):  
Natasha T. Strande ◽  
Juan Carvajal-Garcia ◽  
Ryan A. Hallett ◽  
Crystal A. Waters ◽  
Steven A. Roberts ◽  
...  
Keyword(s):  
Lyase Activity ◽  
Ap Lyase

scholarly journals A covalent protein–DNA 5′-product adduct is generated following AP lyase activity of human ALKBH1 (AlkB homologue 1)

2013 ◽  
Vol 452 (3) ◽  
pp. 509-518 ◽  
Author(s):  
Tina A. Müller ◽  
Megan M. Andrzejak ◽  
Robert P. Hausinger
Keyword(s):  
Dna Adduct ◽  
Covalent Attachment ◽  
Main Site ◽  
Chemical Mechanisms ◽  
Covalent Adduct ◽  
Lysine Residues ◽  
Lyase Activity ◽  
Ap Lyase ◽  
Covalent Intermediate

ALKBH1 (AlkB homologue 1) is a mammalian AlkB (2-oxoglutarate-dependent dioxygenase) homologue that possesses AP (abasic or apurinic/apyrimidinic) lyase activity. The AP lyase reaction is catalysed by imine formation with an active site lysine residue, and a covalent intermediate can be trapped in the presence of NaBH4. Surprisingly, ALKBH1 also forms a stable protein–DNA adduct in the absence of a reducing agent. Experiments with different substrates demonstrated that the protein covalently binds to the 5′ DNA product, i.e. the fragment containing an α,β-unsaturated aldehyde. The N-terminal domain of ALKBH1 was identified as the main site of linkage with DNA. By contrast, mutagenesis studies suggest that the primary catalytic residue forming the imine linkage is Lys133, with Lys154 and other lysine residues in this region serving in opportunistic roles. These findings confirm the classification of ALKBH1 as an AP lyase, identify the primary and a secondary lysine residues involved in the lyase reaction, and demonstrate that the protein forms a covalent adduct with the 5′ DNA product. We propose two plausible chemical mechanisms to account for the covalent attachment.

scholarly journals Molecular Cloning and Functional Analysis of the MutY Homolog of Deinococcus radiodurans

2001 ◽  
Vol 183 (21) ◽  
pp. 6151-6158 ◽  
Author(s):  
Xianghong Li ◽  
A-Lien Lu
Keyword(s):  
Oxidative Stress ◽  
Mutation Frequency ◽  
Deinococcus Radiodurans ◽  
Binding Affinities ◽  
Dna Complexes ◽  
E Coli ◽  
Binding Preference ◽  
Lyase Activity ◽  
Adenine Glycosylase ◽  
And Oxidative Stress

ABSTRACT The mutY homolog gene (mutY Dr) from Deinococcus radiodurans encodes a 39.4-kDa protein consisting of 363 amino acids that displays 35% identity to the Escherichia coli MutY (MutYEc) protein. Expressed MutYDr is able to complement E. coli mutYmutants but not mutM mutants to reduce the mutation frequency. The glycosylase and binding activities of MutYDr with an A/G-containing substrate are more sensitive to high salt and EDTA concentrations than the activities with an A/7,8-dihydro-8-oxoguanine (GO)-containing substrate are. Like the MutYEc protein, purified recombinant MutYDrexpressed in E. coli has adenine glycosylase activity with A/G, A/C, and A/GO mismatches and weak guanine glycosylase activity with a G/GO mismatch. However, MutYDrexhibits limited apurinic/apyrimidinic lyase activity and can form only weak covalent protein-DNA complexes in the presence of sodium borohydride. This may be due to an arginine residue that is present in MutYDr at the position corresponding to the position of MutYEc Lys142, which forms the Schiff base with DNA. The kinetic parameters of MutYDr are similar to those of MutYEc. Although MutYDr has similar substrate specificity and a binding preference for an A/GO mismatch over an A/G mismatch, as MutYEc does, the binding affinities for both mismatches are slightly lower for MutYDr than for MutYEc. Thus, MutYDr can protect the cell from GO mutational effects caused by ionizing radiation and oxidative stress.

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