β-d-glucosyl-hydroxymethyluracil: A novel modified base present in the DNA of the parasitic protozoan T. brucei

Gommers-Ampt, Janet H.; Leeuwen, Fred van; Beer, Antonius L. J. De; Vliegenthart, Johannes F.G.; Dizdaroğlu, Miral; Kowalak, Jeffrey A.; Crain, Pamela F.; Borst, Piet

doi:10.1016/0092-8674(93)90322-h

Cited by 200 publications

(125 citation statements)

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“…Up to one percent of the thymine in the telomeric and subtelomeric repeats of nuclear DNA in the bloodstream form of T. brucei is replaced with β-D-glucosyl-hydroxymethyluracil (base J) but no modification is found in the insect form of the parasite where antigenic variation is absent. 41,42 The modified base is thought to induce chromatin compaction and thereby stabilize the chromosome from rearrangements within these repetitive sequences. 43,44 Moreover, base J is suggested to play a role in gene silencing related to the process of antigenic variation.…”

Section: Polynucleotide Hydroxylases 41 Thymidine Hydroxylase Of Promentioning

confidence: 99%

FeII/α-ketoglutarate hydroxylases involved in nucleobase, nucleoside, nucleotide, and chromatin metabolism

2008

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Fe II /α-ketoglutarate-dependent hydroxylases uniformly possess a double-stranded β-helix fold with two conserved histidines and one carboxylate coordinating their mononuclear ferrous ions. Oxidative decomposition of the α-keto acid is proposed to generate a ferryl-oxo intermediate capable of hydroxylating unactivated carbon atoms in a myriad of substrates. This perspective focuses on a subgroup of these enzymes that are involved in pyrimidine salvage, purine decomposition, nucleoside and nucleotide hydroxylation, DNA/RNA repair, and chromatin modification. The varied reaction schemes are presented, and selected structural and kinetic information is summarized. IntroductionFerrous ion and α-ketoglutarate (αKG)-dependent dioxygenases comprise an enzyme superfamily with members present in animals, plants, protists, fungi, bacteria and even viruses. Most representatives of this large group of enzymes couple the oxidative decarboxylation of αKG to various hydroxylation reactions ( Fig. 1), but others are capable of catalyzing desaturation, epoxidation, halogenation, ring formation, or ring expansion reactions, which allows them to fill a wide variety of biological roles including antibiotic biosynthesis, hypoxic sensing, DNA repair, and various types of metabolite transformations. [1][2][3] Not surprisingly, the primary substrates also are diverse and include proteins, lipids, nucleic acids, and a myriad of small molecules that can be used for synthesis or undergo degradation.The sequences of Fe II /αKG dioxygenases show little overall identity, yet the diverse reactions all are catalyzed by proteins with the same core fold and use similar chemistries. 4 The hallmark of these enzymes is their use of Fe II to activate molecular oxygen according to a mechanism (Scheme 1) first proposed by Hanauske-Abel and Günzler in 1982 for prolyl 4-hydroxylase. 5 (A) In the absence of substrates, a "facial triad" (usually two His plus one Asp or Glu occurring in a His-X-Asp/Glu-X n -His motif) weakly bind one face of the hexacoordinate metal. 6,7 (B) The αKG co-substrate displaces two waters and coordinates Fe II in a bidentate fashion with its C1 carboxyl and C2 keto oxygens. The binding of αKG is Correspondence to: Robert P. Hausinger, hausinge@msu.edu. NIH Public Access Author ManuscriptDalton Trans. Author manuscript; available in PMC 2010 July 20. This perspective focuses on the modification of nucleobases, nucleosides, nucleotides, and chromatin by the Fe II and αKG dependent hydroxylases (Table 1). We discuss the degradation and salvage of free bases by the fungal enzymes thymine 7-hydroxylase and xanthine hydroxylase. Next, nucleoside hydroxylases are briefly described. We then examine the developmental modification of DNA in kinetoplastid protozoans and summarize evidence that the JBP1 and JBP2 proteins possess thymidine hydroxylase activity. The oxidative repair of DNA by Escherichia coli AlkB is discussed, and the properties of mammalian homologues are summarized. Finally, we present emerging evidence pointing to...

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Section: Polynucleotide Hydroxylases 41 Thymidine Hydroxylase Of Promentioning

confidence: 99%

FeII/α-ketoglutarate hydroxylases involved in nucleobase, nucleoside, nucleotide, and chromatin metabolism

2008

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“…The modifications vary from a simple addition of a methyl or hydroxyl group to the 'very unusual' (or hypermodified) bases such as glucosylated hydroxymethylcytosine in T-even bacteriophage DNA (Warren, 1980) and β-D-glucosyl-hydroxymethyluracil (or J) identified in the unicellular eukaryote Trypanosoma brucei (Gommers-Ampt et al, 1993). The modified bases serve various important biological functions, such as protection against degradation by nucleases and regulation of gene expression.…”

Section: Introductionmentioning

confidence: 99%

Crystal structure of deoxycytidylate hydroxymethylase from bacteriophage T4, a component of the deoxyribonucleoside triphosphate-synthesizing complex

1999

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Bacteriophage T4 deoxycytidylate hydroxymethylase (EC 2.1.2.8), a homodimer of 246-residue subunits, catalyzes hydroxymethylation of the cytosine base in deoxycytidylate (dCMP) to produce 5-hydroxymethyldCMP. It forms part of a phage DNA protection system and appears to function in vivo as a component of a multienzyme complex called deoxyribonucleoside triphosphate (dNTP) synthetase. We have determined its crystal structure in the presence of the substrate dCMP at 1.6 Å resolution. The structure reveals a subunit fold and a dimerization pattern in common with thymidylate synthases, despite low (~20%) sequence identity. Among the residues that form the dCMP binding site, those interacting with the sugar and phosphate are arranged in a configuration similar to the deoxyuridylate binding site of thymidylate synthases. However, the residues interacting directly or indirectly with the cytosine base show a more divergent structure and the presumed folate cofactor binding site is more open. Our structure reveals a water molecule properly positioned near C-6 of cytosine to add to the C-7 methylene intermediate during the last step of hydroxymethylation. On the basis of sequence comparison and crystal packing analysis, a hypothetical model for the interaction between T4 deoxycytidylate hydroxymethylase and T4 thymidylate synthase in the dNTP-synthesizing complex has been built. Keywords: bacteriophage T4/deoxycytidylate hydroxymethylase/deoxyuridylate hydroxymethylase/ dNTP-synthesizing complex/thymidylate synthase

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“…In the DNA of kinetoplastid flagellates, a fraction of thymine is replaced by the modified base β-D-glucosyl-hydroxymethyluracil, called J [1][2][3]. This hyper-modified base consists of the attachment of a bulky glucose moiety to the thymine base such that it extends into the major groove of the DNA helix.…”

Section: Introductionmentioning

confidence: 99%

JBP2, a SWI2/SNF2-like protein, regulates de novo telomeric DNA glycosylation in bloodstream form Trypanosoma brucei

Kieft

Brand

Ekanayake

et al. 2007

Molecular and Biochemical Parasitology

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Synthesis of the modified thymine base, β-D-glucosyl-hydroxymethyluracil or J, within telomeric DNA of Trypanosoma brucei correlates with the bloodstream-form specific epigenetic silencing of telomeric variant surface glycoprotein genes involved in antigenic variation. In order to analyze the function of base J in the regulation of antigenic variation, we are characterizing the regulatory mechanism of J biosynthesis. We have recently proposed a model in which chromatin remodeling by a SWI2/SNF2-like protein (JBP2) regulates the developmental and de-novo site-specific localization of J synthesis within bloodstream-form trypanosome DNA. Consistent with this model, we now show that JBP2 (−/−) bloodstream-form trypanosomes contain 5-fold less base J and are unable to stimulate de-novo J synthesis in newly generated telomeric arrays.

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β-d-glucosyl-hydroxymethyluracil: A novel modified base present in the DNA of the parasitic protozoan T. brucei

Cited by 200 publications

References 29 publications

FeII/α-ketoglutarate hydroxylases involved in nucleobase, nucleoside, nucleotide, and chromatin metabolism

FeII/α-ketoglutarate hydroxylases involved in nucleobase, nucleoside, nucleotide, and chromatin metabolism

Crystal structure of deoxycytidylate hydroxymethylase from bacteriophage T4, a component of the deoxyribonucleoside triphosphate-synthesizing complex

JBP2, a SWI2/SNF2-like protein, regulates de novo telomeric DNA glycosylation in bloodstream form Trypanosoma brucei

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