Structure and function of a novel purine specific nucleoside hydrolase from Trypanosoma vivax11Edited by R. Huber

Versées, Wim; Decanniere, Klaas; Pellé, Roger; Depoorter, J.; Brosens, Elke; Parkin, David W.; Steyaert, Jan

doi:10.1006/jmbi.2001.4548

Cited by 92 publications

(147 citation statements)

References 65 publications

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“…Production of the Mutant IAG-NHs-The T. vivax inosine-adenosineguanosine preferring nucleoside hydrolase (iagnh) open reading frame cloned into the BamHI-PstI restriction sites of the pQE-30 plasmid (Qiagen) was used as the template for all in vitro mutagenic studies (1). Mutants were created using a primer-extension overlap PCR technique (18).…”

Section: Methodsmentioning

confidence: 99%

“…Kinetic Analysis of Substrates-The kinetic properties of the IAG-NH for the substrates xanthosine and purine riboside were determined with the reducing sugar assay as described previously (1,9). Hydrolysis of adenosine, 2Ј-deoxyadenosine, and 5Ј-deoxyadenosine was monitored spectrophotometrically using the difference in absorption between the nucleoside and the purine base.…”

Section: Methodsmentioning

confidence: 99%

“…For a detailed description of the overall fold and the active site architecture of the wild type IAG-NH from T. vivax we refer to Ref. 1.…”

Section: Effect Of Ph On the Kinetic Constants Of The Iag-nucleoside mentioning

confidence: 99%

“…Nucleoside hydrolases (NHs) 1 catalyze the irreversible hydrolysis of the N-glycosidic bond in ribonucleosides following the reaction scheme: ␤-purine (or pyrimidine) nucleoside ϩ H 2 O 3 purine (or pyrimidine) base ϩ ribose. The NHs are members of a broader class of N-ribohydrolases and transferases including the clinically important purine-nucleoside phosphorylases (2) and phosphoribosyl transferases (3,4).…”

mentioning

confidence: 99%

“…Since neither nucleoside hydrolase activity, nor the encoding genes, have been found in mammals they constitute an attractive target for drug design against a variety of pathogens. On the basis of their substrate specificity the nucleoside hydrolases have been divided into three subclasses: the base aspecific inosine-uridine preferring nucleoside hydrolases (IU-NH) (8), the purine-specific inosine-adenosineguanosine preferring nucleoside hydrolases (IAG-NH) (1,9), and the 6-oxo-purine specific inosine-guanosine preferring nucleoside hydrolases (IG-NH) (10).…”

mentioning

confidence: 99%

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Enzyme-Substrate Interactions in the Purine-specific Nucleoside Hydrolase from Trypanosoma vivax

Versées

Decanniere

Holsbeke

et al. 2002

Journal of Biological Chemistry

131

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Nucleoside hydrolases are key enzymes in the purine salvage pathway of Trypanosomatidae and are considered as targets for drug design. We previously reported the first x-ray structure of an inosine-adenosineguanosine preferring nucleoside hydrolase (IAG-NH) from Trypanosoma vivax (1). Here we report the 2.0-Å crystal structure of the slow D10A mutant in complex with the inhibitor 3-deaza-adenosine and the 1.6-Å crystal structure of the same enzyme in complex with a genuine substrate inosine. The enzyme-substrate complex shows the substrate bound to the enzyme in a different conformation from 3-deaza-adenosine and provides a snapshot along the reaction coordinate of the enzyme-catalyzed reaction. The chemical groups on the substrate important for binding and catalysis are mapped. The 2-OH, 3-OH, and 5-OH contribute 4.6, 7.5, and 5.4 kcal/mol to k cat /K m , respectively. Specific interactions with the exocyclic groups on the purine ring are not required for catalysis. Site-directed mutagenesis indicates that the purine specificity of the IAG-NHs is imposed by a parallel aromatic stacking interaction involving Trp 83 and Trp 260. The pH profiles of k cat and k cat /K m indicate the existence of one or more proton donors, possibly involved in leaving group activation. However, mutagenesis of the active site residues around the nucleoside base and an alanine scan of a flexible loop near the active site fail to identify this general acid. The parallel aromatic stacking seems to provide the most likely alternative mechanism for leaving group activation.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

“…For a detailed description of the overall fold and the active site architecture of the wild type IAG-NH from T. vivax we refer to Ref. 1.…”

Section: Effect Of Ph On the Kinetic Constants Of The Iag-nucleoside mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Enzyme-Substrate Interactions in the Purine-specific Nucleoside Hydrolase from Trypanosoma vivax

Versées

Decanniere

Holsbeke

et al. 2002

Journal of Biological Chemistry

131

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Synthesis of Bicyclic N‐Arylmethyl‐Substituted Iminoribitol Derivatives as Selective Nucleoside Hydrolase Inhibitors

et al. 2009

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The purine metabolism of Trypanosoma and Leishmania spp. provides a good target in the search for new selective drugs. Bicyclic N-arylmethyl-substituted iminoribitols were developed as inhibitors of T. vivax nucleoside hydrolase, a key enzyme of the purine salvage pathway. The obtained results and structure-activity data confirmed our model for inhibitor binding with a hydrogen bond between a nitrogen atom of the nucleobase mimetic and the protonated Asp40 from the enzyme. This interaction depends on an optimal pK(a) value, which can be influenced by the electronic properties of the substituents. These compounds are potent, selective inhibitors of nucleoside hydrolase and are inactive toward human nucleoside phosphorylase.

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Structural and biochemical characterization of the nucleoside hydrolase from C. elegans reveals the role of two active site cysteine residues in catalysis

2017

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Nucleoside hydrolases (NHs) catalyze the hydrolysis of the N-glycoside bond in ribonucleosides and are found in all three domains of life. Although in parasitic protozoa a role in purine salvage has been well established, their precise function in bacteria and higher eukaryotes is still largely unknown. NHs have been classified into three homology groups based on the conservation of active site residues. While many structures are available of representatives of group I and II, structural information for group III NHs is lacking. Here, we report the first crystal structure of a purine-specific nucleoside hydrolase belonging to homology group III from the nematode Caenorhabditis elegans (CeNH) to 1.65Å resolution. In contrast to dimeric purine-specific NHs from group II, CeNH is a homotetramer. A cysteine residue that characterizes group III NHs (Cys253) structurally aligns with the catalytic histidine and tryptophan residues of group I and group II enzymes, respectively. Moreover, a second cysteine (Cys42) points into the active site of CeNH. Substrate docking shows that both cysteine residues are appropriately positioned to interact with the purine ring. Site-directed mutagenesis and kinetic analysis proposes a catalytic role for both cysteines residues, with Cys253 playing the most prominent role in leaving group activation.

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Structure and function of a novel purine specific nucleoside hydrolase from Trypanosoma vivax11Edited by R. Huber

Cited by 92 publications

References 65 publications

Enzyme-Substrate Interactions in the Purine-specific Nucleoside Hydrolase from Trypanosoma vivax

Enzyme-Substrate Interactions in the Purine-specific Nucleoside Hydrolase from Trypanosoma vivax

Synthesis of Bicyclic N‐Arylmethyl‐Substituted Iminoribitol Derivatives as Selective Nucleoside Hydrolase Inhibitors

Structural and biochemical characterization of the nucleoside hydrolase from C. elegans reveals the role of two active site cysteine residues in catalysis

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