Characterization of the apurinic endonuclease activity of Drosophila Rrp1

Nugent, Mark; Huang, Shao-Liang; Sander, Miriam

doi:10.1021/bi00093a023

Cited by 13 publications

(12 citation statements)

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“…Purified human Ref-1 used as a control was the generous gift of Steven Xanthodakis (Roche Biomedical, Nutley, N.J.). Additional AP endonuclease assays were performed in the laboratory of Miriam Sander (NIEHS, Research Triangle Park, N.C.) by using a synthetic oligonucleotide substrate (25). Exonuclease assays were performed on linearized plasmid DNA with E. coli ExoIII (Promega Corporation) as a positive control (3).…”

Section: Methodsmentioning

confidence: 99%

The nucleotide sequence of the Pseudomonas aeruginosa pyrE-crc-rph region and the purification of the crc gene product

et al. 1996

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The gene (crc) responsible for catabolite repression control in Pseudomonas aeruginosa has been cloned and sequenced. Flanking the crc gene are genes encoding orotate phosphoribosyl transferase (pyrE) and RNase PH (rph). New crc mutants were constructed by disruption of the wild-type crc gene. The crc gene encodes an open reading frame of 259 amino acids with homology to the apurinic/apyrimidinic endonuclease family of DNA repair enzymes. However, crc mutants do not have a DNA repair phenotype, nor can the crc gene complement Escherichia coli DNA repair-deficient strains. The crc gene product was overexpressed in both P. aeruginosa and in E. coli, and the Crc protein was purified from both. The purified Crc proteins show neither apurinic/ apyrimidinic endonuclease nor exonuclease activity. Antibody to the purified Crc protein reacted with proteins of similar size in crude extracts from Pseudomonas putida and Pseudomonas fluorescens, suggesting a common mechanism of catabolite repression in these three species.The genus Pseudomonas is noteworthy for its diversity in habitat and physiology. Some Pseudomonas strains are able to use over 100 organic compounds as the sole or principal source of carbon. Pseudomonas aeruginosa can utilize at least 80 different organic compounds (41). A mechanism of catabolite repression control exists in these organisms (18) which prevents them from wasting energy maintaining the enzymes for all these catabolic pathways and ensures the preferential utilization of the most efficient source of carbon and energy. Such a regulatory mechanism has also been identified in the enteric bacteria (20) and in Bacillus spp. (6).In the enteric bacteria, the molecular mechanism of catabolite repression control involves a catabolite activator protein (Cap) which, when bound to cyclic AMP (cAMP), interacts with promoter regions of regulated genes to facilitate the binding of RNA polymerase, thereby initiating transcription (20). In the presence of glucose, the cAMP pool is lowered, Cap is not bound, and the regulated genes are not transcribed (i.e., they are repressed). The effect of glucose on cAMP pools is mediated by components of the phosphoenolpyruvate phosphotransferase system which also serve to activate adenylate cyclase (33). Since the initial identification of these regulatory components, catabolite repression has proven to be a global mechanism in the enteric bacteria, affecting at least 28 separate promoters which regulate biosynthetic as well as catabolic operons (7). It also has been found to act as a negative regulator as well as a positive regulator (1,7,23,24).In Bacillus spp. catabolite repression is not mediated by glucose, nor is cAMP involved (6,37,40). Genetic evidence has implicated the catabolite control protein (CcpA), a member of the GalR family of repressor proteins (6), Hpr, a component of the phosphoenolpyruvate phosphotransferase system (9), and a cis-acting DNA sequence (CRE) (15,48). Recent work has shown that HPr specifically phosphorylated at Ser-46 forms a complex wit...

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Section: Methodsmentioning

confidence: 99%

The nucleotide sequence of the Pseudomonas aeruginosa pyrE-crc-rph region and the purification of the crc gene product

et al. 1996

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“…The compact C-terminal part (251 aa long; Figure 1B) was later shown to bear homology with EEP superfamily AP endonucleases [174]. It catalyzes cleavage of AP sites, resection of blunt or recessed 3′-ends in a DNA duplex, possesses 3′-phosphatase and 3′phosphodiesterase activities on recessed damaged 3′-ends [175][176][177][178] and complements the oxidation-and alkylation-sensitive phenotype of AP endonuclease-null E. coli [179], showing all attributes of a functional AP endonuclease participating in BER. Although no structural information on Rrp1 is available, limited proteolysis combined with circular dichroism and sedimentation velocity analysis indicate that the N-tail (426 aa) is mostly unstructured [180].…”

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confidence: 99%

Reading Targeted DNA Damage in the Active Demethylation Pathway: Role of Accessory Domains of Eukaryotic AP Endonucleases and Thymine-DNA Glycosylases

Popov

Grin

Dvornikova

et al. 2020

Journal of Molecular Biology

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Base excision DNA repair (BER) is an important process used by all living organisms to remove non-bulky lesions from DNA. BER is usually initiated by DNA glycosylases that excise a damaged base leaving an apurinic/apyrimidinic (AP) site, an AP endonuclease then cuts DNA at the AP site, and the repair is completed by correct nucleotide insertion, end processing, and nick ligation. It has emerged recently that the BER machinery, in addition to genome protection, is crucial for active epigenetic demethylation in vertebrates. This pathway is initiated by TET dioxygenases that oxidize the regulatory 5-methylcytosine, and the oxidation products are treated as substrates for BER. T:G mismatch-specific thymine-DNA glycosylase (TDG) and AP endonuclease 1 (APE1) catalyze the first two steps in BER-dependent active demethylation. In addition to the well-structured catalytic domains, these enzymes possess long tails that are structurally uncharacterized but involved in multiple interactions and regulatory functions. In this review, we describe known roles of the tails in TDG and APE1, discuss the importance of order and disorder in their structure and consider the evolutionary aspects of these accessory protein regions. We also propose that the tails may be important for the enzymes' oligomerization on DNA, an aspect of their function that only recently gained attention.

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“…Drosophila Rrpl is a well-characterized AP endonuclease that is enzymatically similar to E. coli exonuclease III (9). Its nucleolytic functions, encoded by the C-terminal third of Rrpl,…”

mentioning

confidence: 99%

“…include double-stranded DNA 3'-exonuclease, AP endonuclease, 3'-phosphodiesterase, and 3'-phosphatase (9)(10)(11) (17,18). This fact has been exploited for a wide range of different applications, including the study of developmental processes (20)(21)(22)(23) and cell viability (24,25).…”

mentioning

confidence: 99%

Overexpression of a Rrp1 transgene reduces the somatic mutation and recombination frequency induced by oxidative DNA damage in Drosophila melanogaster.

Szakmary¹,

Huang²,

Chang³

et al. 1996

Proc. Natl. Acad. Sci. U.S.A.

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Recombination repair protein 1 (Rrpl) includes a C-terminal region homologous to several DNA repair proteins, including Escherichia coli exonuclease III and human APE, that repair oxidative and alkylation damage to DNA. The nuclease activities of Rrpl include apurinic/apyrimidinic endonuclease, 3'-phosphodiesterase, 3'-phosphatase, and 3'-exonuclease. As shown previously, the C-terminal nuclease region of Rrpl is sufficient to repair oxidative-and alkylationinduced DNA damage in repair-deficient E. coli mutants. DNA strand-transfer and single-stranded DNA renaturation activities are associated with the unique N-terminal region of Rrpl, which suggests possible additional functions that include recombinational repair or homologous recombination. By using the Drosophila w/w+ mosaic eye system, which detects loss of heterozygosity as changes in eye pigmentation, somatic mutation and recombination frequencies were determined in transgenic flies overexpressing wild-type Rrpl protein from a heat-shock-inducible transgene. A large decrease in mosaic clone frequency is observed when Rrpl overexpression precedes treatment with y-rays, bleomycin, or paraquat. In contrast, Rrpl overexpression does not alter the spot frequency after treatment with the alkylating agents methyl methanesulfonate or methyl nitrosourea. A reduction in mosaic clone frequency depends on the expression of the Rrpl transgene and on the nature of the induced DNA damage. These data suggest a lesion-specific involvement ofRrpl in the repair of oxidative DNA damage.DNA repair enzymes are ubiquitous in all organisms studied and display a large range of functional specificities, including both exquisitely specialized and very broad selectivities (1-4). These enzymes allow cells and organisms to preserve genomic integrity, to prevent the accumulation of deleterious mutations, and, in some cases, to survive lethal challenges.A major class of DNA damage is caused by reactive oxygen species that lead to altered DNA bases and to both singlestranded DNA (ssDNA) and double-stranded DNA breaks (5, 6). Oxidative damage is thought to be mutagenic, clastogenic, and carcinogenic and may contribute to the aging process. The general pathway known as base excision repair plays a major role in repair of oxidative DNA damage. One crucial enzyme involved in this pathway is the class II apurinic/apyrimidinic (AP) endonuclease, of which two major families are known, the exonuclease III family and the endonuclease IV family. Mutants of either Escherichia coli exonuclease III or endonuclease IV (or its yeast homologue APN1) are sensitive to oxidative and alkylation DNA damage and show increased spontaneous mutation rates (7,8).Drosophila Rrpl is a well-characterized AP endonuclease that is enzymatically similar to E. coli exonuclease III (9). Its nucleolytic functions, encoded by the C-terminal third of Rrpl,The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with ...

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Characterization of the apurinic endonuclease activity of Drosophila Rrp1

Cited by 13 publications

References 29 publications

The nucleotide sequence of the Pseudomonas aeruginosa pyrE-crc-rph region and the purification of the crc gene product

The nucleotide sequence of the Pseudomonas aeruginosa pyrE-crc-rph region and the purification of the crc gene product

Reading Targeted DNA Damage in the Active Demethylation Pathway: Role of Accessory Domains of Eukaryotic AP Endonucleases and Thymine-DNA Glycosylases

Overexpression of a Rrp1 transgene reduces the somatic mutation and recombination frequency induced by oxidative DNA damage in Drosophila melanogaster.

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