Structures of dCTP Deaminase from Escherichia coli with Bound Substrate and Product

Johansson, E.; Fano, Mathias; Bynck, J.H.; Neuhard, Jan; Larsen, Sine; Sigurskjold, Bent W.; Christensen, Ulla; Willemoës, Martin

doi:10.1074/jbc.m409534200

Cited by 35 publications

(30 citation statements)

References 38 publications

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“…However, in recent structural investigations, some members of the dUTPase trimeric superfamily were shown to lack this arm-swapping interaction (Figure 6). Namely, the bifunctional dCTP-deaminase-dUTPase from Archea, the Plasmodium falciparum dUTPase, as well as the more distantly related E.coli dCTP-deaminase proteins all display homotrimeric organization with similar active site architecture as most dUTPases, but arm-swapping is not present in any of these structures (Figure 6) (31–33). The present investigation on M-PMV dUTPase clearly demonstrates that despite lack of arm-swapping, the C-terminal segment retains a major role in active site architecture: the truncated mutant is characterized by a 10 5 -fold decrease in catalytic efficiency ( k cat / K M ).…”

Section: Discussionmentioning

confidence: 99%

Flexible segments modulate co-folding of dUTPase and nucleocapsid proteins

Németh-Pongrácz¹,

Barábas²,

Fuxreiter³

et al. 2006

Nucleic Acids Research

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The homotrimeric fusion protein nucleocapsid (NC)-dUTPase combines domains that participate in RNA/DNA folding, reverse transcription, and DNA repair in Mason-Pfizer monkey betaretrovirus infected cells. The structural organization of the fusion protein remained obscured by the N- and C-terminal flexible segments of dUTPase and the linker region connecting the two domains that are invisible in electron density maps. Small-angle X-ray scattering reveals that upon oligonucleotide binding the NC domains adopt the trimeric symmetry of dUTPase. High-resolution X-ray structures together with molecular modeling indicate that fusion with NC domains dramatically alters the conformation of the flexible C-terminus by perturbing the orientation of a critical β-strand. Consequently, the C-terminal segment is capable of double backing upon the active site of its own monomer and stabilized by non-covalent interactions formed with the N-terminal segment. This co-folding of the dUTPase terminal segments, not observable in other homologous enzymes, is due to the presence of the fused NC domain. Structural and genomic advantages of fusing the NC domain to a shortened dUTPase in betaretroviruses and the possible physiological consequences are envisaged.

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Section: Discussionmentioning

confidence: 99%

Flexible segments modulate co-folding of dUTPase and nucleocapsid proteins

Németh-Pongrácz¹,

Barábas²,

Fuxreiter³

et al. 2006

Nucleic Acids Research

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“…The asymmetric unit contains 6 molecules arranged in two trimers. The homotrimeric form is a common feature of the enzyme family (18). The different subunits align to each other with root mean square deviations (RMSDs) ranging from 0.169 Å (subunits A and B) to 0.309 Å (subunits A and C).…”

Section: Resultsmentioning

confidence: 99%

“…Enteric bacteria synthesize an enzyme with dCTP deaminase activity that converts dCTP into dUTP (EC 3.5.4.13), which accounts for 70 to 80% of the dUMP production (15). The dCTP deaminase is a homotrimeric enzyme structurally related to the trimeric dUTPase and part of the dCTP deaminase/dUTPase superfamily (16)(17)(18). The genomes of many Gram-positive bacteria and all eukaryotes encode a dCMP deaminase (EC 3.5.4.12) that shares no resemblance to the enteric dCTP deaminase (14,(19)(20)(21)(22)(23).…”

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confidence: 99%

“…The genomes of many Gram-positive bacteria and all eukaryotes encode a dCMP deaminase (EC 3.5.4.12) that shares no resemblance to the enteric dCTP deaminase (14,(19)(20)(21)(22)(23). The dCTP deaminase catalyzes dCTP deamination solely by way of amino acid residues present in the active site (16,18), while the dCMP deaminase, a hexameric enzyme which is related to other cytosine deaminating enzymes, such as cytosine deaminase (24) and cytidine deaminase (25)(26)(27)(28)(29), binds a zinc ion that directly participates in the catalytic mechanism (14,19,20,30). One feature shared by the two enzymes, dCTP and dCMP deaminase, is that they are both feedback inhibited by dTTP (17,19,(31)(32)(33)(34)(35)(36)(37)(38).…”

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confidence: 99%

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Bacillus halodurans Strain C125 Encodes and Synthesizes Enzymes from Both Known Pathways To Form dUMP Directly from Cytosine Deoxyribonucleotides

Oehlenschlæger

Løvgreen

Reinauer

et al. 2015

Appl Environ Microbiol

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Analysis of the genome of Bacillus halodurans strain C125 indicated that two pathways leading from a cytosine deoxyribonucleotide to dUMP, used for dTMP synthesis, were encoded by the genome of the bacterium. The genes that were responsible, the comEB gene and the dcdB gene, encoding dCMP deaminase and the bifunctional dCTP deaminase:dUTPase (DCD:DUT), respectively, were both shown to be expressed in B. halodurans, and both genes were subject to repression by the nucleosides thymidine and deoxycytidine. The latter nucleoside presumably exerts its repression after deamination by cytidine deaminase. Both comEB and dcdB were cloned, overexpressed in Escherichia coli, and purified to homogeneity. Both enzymes were active and displayed the expected regulatory properties: activation by dCTP for dCMP deaminase and dTTP inhibition for both enzymes. Structurally, the B. halodurans enzyme resembled the Mycobacterium tuberculosis enzyme the most. An investigation of sequenced genomes from other species of the genus Bacillus revealed that not only the genome of B. halodurans but also the genomes of Bacillus pseudofirmus, Bacillus thuringiensis, Bacillus hemicellulosilyticus, Bacillus marmarensis, Bacillus cereus, and Bacillus megaterium encode both the dCMP deaminase and the DCD:DUT enzymes. In addition, eight dcdB homologs from Bacillus species within the genus for which the whole genome has not yet been sequenced were registered in the NCBI Entrez database. The biosynthetic route to the nucleotide dUMP, the precursor of dTTP, as shown in Fig. 1, differs between organisms and sets it apart from the rest of nucleotide metabolism (1). This is in agreement with the general belief that uracil was the original DNA base-pairing partner for adenine and that the biosynthesis of dTTP from dUMP arose later in evolution (2). The deoxyribonucleotides dGTP, dATP, and dCTP are all produced in a pathway that goes through the multisubstrate enzyme ribonucleotide reductase. This enzyme is highly regulated in order to maintain the balance of deoxyribonucleotide pools (3-5). dUTP is also formed by ribonucleotide reductase, but because of the toxicity of this nucleotide (6, 7), all organisms express an enzyme with dUTPase activity that hydrolyzes dUTP to dUMP (8).The direct formation of dUMP via ribonucleotide reductase (Fig. 1) is not an efficient process, as the ribonucleotide reductase has a reduced affinity for UDP compared to the affinities of three other canonical ribonucleotides (9-12). Instead, most of the dUMP arises from deamination of cytosine deoxyribonucleotides (13-15). Enteric bacteria synthesize an enzyme with dCTP deaminase activity that converts dCTP into dUTP (EC 3.5.4.13), which accounts for 70 to 80% of the dUMP production (15). The dCTP deaminase is a homotrimeric enzyme structurally related to the trimeric dUTPase and part of the dCTP deaminase/dUTPase superfamily (16-18). The genomes of many Gram-positive bacteria and all eukaryotes encode a dCMP deaminase (EC 3.5.4.12) that shares no resemblance to the enteric dCTP de...

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“…Such modifications play a central role in RNA editing, which is critical for generating the appropriate anti-codon sequences for decoding the genetic code, modification of the sequences of microRNA and other transcripts and alteration of the reading frames in mRNAs, defense against viruses via hypermutation-based inactivation, and somatic hypermutation or class switching of antigen receptor genes in vertebrates (1,5–8). In addition to the deaminase superfamily, deamination of standalone bases is also catalyzed by structurally unrelated amidohydrolases that display other protein folds, such as the CodA-like cytosine deaminases and Amd1-like AMP deaminases with a TIM Barrel fold (9) and Escherichia coli Dcd-like dCTP deaminases with the dUTPase fold (10). However, currently, only members of the deaminase superfamily have been implicated in in situ nucleic acid modifications leading to RNA editing or DNA hypermutation, and are accordingly termed nucleic acid deaminases.…”

Section: Introductionmentioning

confidence: 99%

Evolution of the deaminase fold and multiple origins of eukaryotic editing and mutagenic nucleic acid deaminases from bacterial toxin systems

Iyer

Zhang

Rogozin

et al. 2011

Nucleic Acids Research

152

263

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The deaminase-like fold includes, in addition to nucleic acid/nucleotide deaminases, several catalytic domains such as the JAB domain, and others involved in nucleotide and ADP-ribose metabolism. Using sensitive sequence and structural comparison methods, we develop a comprehensive natural classification of the deaminase-like fold and show that its ancestral version was likely to operate on nucleotides or nucleic acids. Consequently, we present evidence that a specific group of JAB domains are likely to possess a DNA repair function, distinct from the previously known deubiquitinating peptidase activity. We also identified numerous previously unknown clades of nucleic acid deaminases. Using inference based on contextual information, we suggest that most of these clades are toxin domains of two distinct classes of bacterial toxin systems, namely polymorphic toxins implicated in bacterial interstrain competition and those that target distantly related cells. Genome context information suggests that these toxins might be delivered via diverse secretory systems, such as Type V, Type VI, PVC and a novel PrsW-like intramembrane peptidase-dependent mechanism. We propose that certain deaminase toxins might be deployed by diverse extracellular and intracellular pathogens as also endosymbionts as effectors targeting nucleic acids of host cells. Our analysis suggests that these toxin deaminases have been acquired by eukaryotes on several independent occasions and recruited as organellar or nucleo-cytoplasmic RNA modifiers, operating on tRNAs, mRNAs and short non-coding RNAs, and also as mutators of hyper-variable genes, viruses and selfish elements. This scenario potentially explains the origin of mutagenic AID/APOBEC-like deaminases, including novel versions from Caenorhabditis, Nematostella and diverse algae and a large class of fast-evolving fungal deaminases. These observations greatly expand the distribution of possible unidentified mutagenic processes catalyzed by nucleic acid deaminases.

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Structures of dCTP Deaminase from Escherichia coli with Bound Substrate and Product

Cited by 35 publications

References 38 publications

Flexible segments modulate co-folding of dUTPase and nucleocapsid proteins

Flexible segments modulate co-folding of dUTPase and nucleocapsid proteins

Bacillus halodurans Strain C125 Encodes and Synthesizes Enzymes from Both Known Pathways To Form dUMP Directly from Cytosine Deoxyribonucleotides

Evolution of the deaminase fold and multiple origins of eukaryotic editing and mutagenic nucleic acid deaminases from bacterial toxin systems

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