Substrate recognition and catalysis by flap endonucleases and related enzymes

Tomlinson, Christopher G.; Atack, John M.; Chapados, Brian R.; Tainer, John A.; Grasby, Jane A.

doi:10.1042/bst0380433

Cited by 42 publications

(58 citation statements)

References 33 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In contrast to the ExoIX subgroup, the carboxylate-rich active site is conserved in the majority of the DNA 5Ј-nuclease superfamily (1). These include enzymes that catalyze structure-specific reactions of an apparently disparate group of nucleic acid substrates including DNA bubbles (xeroderma pigmentosum complementation group G (XPG)) (35), four-way junctions (GEN-1 (xeroderma pigmentosum complementation group G-like gap endonuclease, a putative human Holliday junction resolvase)) (36), as well as flapped, nicked, and 3Ј-overhang DNAs (FEN and EXO1) (2,20,37,38).…”

Section: Discussionmentioning

confidence: 99%

“…An unusual feature of these enzymes is their high density of active site carboxylates that coordinate essential divalent metal ion cofactors. Bacterial and bacteriophage FENs possess eight active site acidic residues, seven of which are conserved in FENs from higher organisms and also in other members of the 5Ј-nuclease superfamily (1)(2)(3)(4). In contrast, a typical metallonuclease active site consists of three or four carboxylates (5).…”

mentioning

confidence: 99%

“…Calcium ions do not support FEN catalysis and inhibit the reactions supported by viable metal cofactors. To establish participation of ions in stabilization of enzyme-substrate complexes, dissociation constants of WT and D201I/D204S-substrate complexes were studied as a function of [ 4 , essential in all life forms, are members of the 5Ј-nuclease superfamily of structure-specific phosphate diesterases that carry out essential divalent metal ion-dependent nucleic acid hydrolysis (1). FENs act upon 5Ј-bifurcated structures known as flaps formed during lagging strand DNA replication and long patch base excision repair.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Neutralizing Mutations of Carboxylates That Bind Metal 2 in T5 Flap Endonuclease Result in an Enzyme That Still Requires Two Metal Ions

Tomlinson

Syson

Sengerová

et al. 2011

Journal of Biological Chemistry

View full text Add to dashboard Cite

Flap endonucleases (FENs) are divalent metal ion-dependent phosphodiesterases. Metallonucleases are often assigned a "two-metal ion mechanism" where both metals contact the scissile phosphate diester. The spacing of the two metal ions observed in T5FEN structures appears to preclude this mechanism. However, the overall reaction catalyzed by wild type (WT) T5FEN requires three Mg 2؉ ions, implying that a third ion is needed during catalysis, and so a two-metal ion mechanism remains possible. , essential in all life forms, are members of the 5Ј-nuclease superfamily of structure-specific phosphate diesterases that carry out essential divalent metal ion-dependent nucleic acid hydrolysis (1). FENs act upon 5Ј-bifurcated structures known as flaps formed during lagging strand DNA replication and long patch base excision repair. Hydrolysis predominantly occurs one nucleotide into the 5Ј-duplex adjoining the site of bifurcation. An unusual feature of these enzymes is their high density of active site carboxylates that coordinate essential divalent metal ion cofactors. Bacterial and bacteriophage FENs possess eight active site acidic residues, seven of which are conserved in FENs from higher organisms and also in other members of the 5Ј-nuclease superfamily (1-4). In contrast, a typical metallonuclease active site consists of three or four carboxylates (5). However, a subclass of FEN paralogues, typified by Escherichia coli ExoIX, exists in eubacteria and appear to lack three of the metal-binding carboxylates (6).Although the most common mechanism suggested for metal-dependent phosphoryl transferases involves two metal ions in contact with the scissile phosphate diester, this is not universally accepted (5, 7-10). Debate about the validity or otherwise of two-metal ion catalysis has focused on several enzymes including flap endonucleases (11-16). Crystallographic studies of substrate-free FENs have demonstrated two active site-bound metal ions (14,(17)(18)(19). Metal ion 1 (M1) occupies a similar position in all structures. However, there is some variation in the position of the site coordinating a second metal ion (M2), often observed in FEN crystal structures. The distance between the M1 and M2 sites varies, and the separations observed are generally much greater than the 4 Å required for both metals to contact the same phosphate diester. For example, in bacteriophage T5FEN, 5 the M1-M2 separation of two Mn 2ϩ ions is 8 Å (Fig. 1A) (17). However, in a substrate-free structure of hFEN1, two magnesium ions are bound with a separation of 3.4 Å in one of three proteins bound to proliferating cell nuclear antigen (Fig. 1B) , 3-(N-morpholino)propane-sulfonic acid. 5 In early literature, T5FEN was referred to as T5 5Ј-3Ј-exonuclease and T5 5Ј-nuclease. In early literature, T4FEN was referred to as referred to also as T4 RNase H.

show abstract

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

Neutralizing Mutations of Carboxylates That Bind Metal 2 in T5 Flap Endonuclease Result in an Enzyme That Still Requires Two Metal Ions

Tomlinson

Syson

Sengerová

et al. 2011

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…Human FEN1, which is the archetypal member of the Rad2 nuclease family (3,4), is located on chromosome 11q12 and consists of two exons and one intron. FEN1 efficiently removed the 5'-flaps generated by Polδ/ε during repair synthesis of long-patch base-excision repair (LP-BER) and removed primers during lagging-strand DNA synthesis and Okazaki fragment processing (3,5,6).…”

Section: Introductionmentioning

confidence: 99%

Flap endonuclease 1 polymorphisms (rs174538 and rs4246215) contribute to an increased cancer risk: Evidence from a meta-analysis

Ren

et al. 2015

Molecular and Clinical Oncology

View full text Add to dashboard Cite

Abstract. Flap endonuclease-1 (FEN1) is a key factor during the maintenance of genomic stability and protection against tumorigenesis. Since the identification of functional polymorphisms of FEN1 (rs174538 and rs4246215), numerous studies have evaluated the association between the two single-nucleotide polymorphisms and cancer risk. To derive a more precise estimation, a meta-analysis was performed on the association between the FEN1 polymorphisms (rs174538 and rs4246215) and cancer risk. Odds ratios (ORs) and 95% confidence intervals (CIs) were used to estimate the strength of the associations. Thirteen case-control studies, including 5,108 cases and 6,382 case-free controls, were identified. For rs174538, individuals with the GG or GA genotype had an increased risk of cancer when compared to the -69AA genotype (AA vs. GG: OR, 1.85; 95% CI, 1.65-2.08; P<0.00001; AA vs. GA: OR, 1.43; 95% CI, 1.27-1.60; P<0.00001; AA vs. GG+GA: OR, 1.28; 95% CI, 1.16-1.42; P<0.00001). For rs4246215, similar results were identified, as the GG or GT genotype was significantly associated with the increased cancer risk when compared to TT (TT vs. GG: OR, 1.71; 95% CI, 1.52-1.92; P<0.00001; TT vs. GT: OR, 1.34; 95% CI, 1.20-1.50; P<0.00001; TT vs. GG+GT: OR, 1.50; 95% CI, 1.35-1.67; P<0.00001). The present meta-analysis indicated that FEN1 rs174538 and rs4246215 polymorphisms may contribute to an increased risk of cancer. IntroductionFlap endonuclease 1 (FEN1) is a versatile, structure specific and multifunctional nuclease involved in DNA replication and repair (1,2). Human FEN1, which is the archetypal member of the Rad2 nuclease family (3,4), is located on chromosome 11q12 and consists of two exons and one intron. FEN1 efficiently removed the 5'-flaps generated by Polδ/ε during repair synthesis of long-patch base-excision repair (LP-BER) and removed primers during lagging-strand DNA synthesis and Okazaki fragment processing (3,5,6). Furthermore, FEN1 can be stimulated to promote apoptotic DNA fragmentation following apoptotic stimuli, acting as a 5' exonuclease (1) and a gap-dependent endonuclease (7,8), as reported via its ability to participate in multiple protein-protein interactions. Thus far, >30 FEN1-interacting proteins have been identified (2). Of these FEN1 interaction partners, proliferating cell nuclear antigen (PCNA), which was initially identified as a replication accessory protein, accompanies FEN1 in all FEN1-involved DNA metabolic pathways except for the apoptotic DNA fragmentation pathway, suggesting a critical role of the FEN1/PCNA interaction in regulating LP-BER (9). A tumor suppressor function for FEN1 has been shown in preclinical models (10-14). Therefore, FEN1 has been considered as a key factor during maintenance of genomic stability and protecting against carcinogenesis.However, being a multifunctional factor, mutation of FEN1 has been suggested to cause genomic instability and predisposition to cancer. The functional impairment of yeast RAD27 (the homolog of mammalian FEN1) leads to a marked increase in th...

show abstract

“…Thus, they bear not only 5 0 exo but also 5 0 flap-endonuclease activities. This activity also exists in monofunctional proteins, such as T4FEN [4]. Eukaryotic polymerases such as human FEN1 and yeast Rad27 also bear this kind of catalytic characteristic [5].…”

Section: Introductionmentioning

confidence: 99%

Effects of cations on small fragment of DNA polymerase I using a novel FRET assay

Zhao¹,

Tang²,

Li³

et al. 2014

ABBS

View full text Add to dashboard Cite

DNA polymerase I (PolI) digested by protease produces a small fragment (SF) containing 5 0 !3 0 exonuclease activity. The 5 0 !3 0 exonuclease activity of polI cleaves the downstream RNA primer strands during DNA replication in vivo. Previous in vitro studies suggested its capability of cleaving duplex from 5 0 terminal and a flap-structure-specific endonuclease activity. From the crystal structures of other nucleases and biochemical data, a two-metal-ion mechanism has been proposed but has not been determined. In this study, we cloned, expressed, and purified the SF protein, and established a novel fluorescence resonance energy transfer (FRET)

show abstract

Substrate recognition and catalysis by flap endonucleases and related enzymes

Cited by 42 publications

References 33 publications

Neutralizing Mutations of Carboxylates That Bind Metal 2 in T5 Flap Endonuclease Result in an Enzyme That Still Requires Two Metal Ions

Neutralizing Mutations of Carboxylates That Bind Metal 2 in T5 Flap Endonuclease Result in an Enzyme That Still Requires Two Metal Ions

Flap endonuclease 1 polymorphisms (rs174538 and rs4246215) contribute to an increased cancer risk: Evidence from a meta-analysis

Effects of cations on small fragment of DNA polymerase I using a novel FRET assay

Contact Info

Product

Resources

About