Purine-specific Nucleoside N-Ribohydrolase from Trypanosoma brucei brucei

Parkin, David W.

doi:10.1074/jbc.271.36.21713

Cited by 90 publications

(121 citation statements)

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“…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%

“…The identity of the two protonated residues is less obvious. In analogy with the IU-NHs, a role as general acids in protonating the purine leaving group, was ascribed to them previously (1,9). For the "low pK a " residue (pK a ϭ 5.6) an aspartate or glutamate with a raised pK a or a histidine residue with reduced pK a could be considered.…”

Section: Is a General Acid Involved In The Enzyme Mechanism?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).…”

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confidence: 99%

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

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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: Is a General Acid Involved In The Enzyme Mechanism?mentioning

confidence: 99%

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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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“…We first investigated the quaternary structure of recombinant TTHA0556 because the purine nucleosidases so far examined are reported to form dimers, tetramers, and pentamers. [6][7][8][9][10][11] An assay mixture containing the purified MBP-fused recombinant (Fig. 2) with a calculated molecular mass of 66.3 kDa (MBP; 42.5 kDa, TTHA0556; 23.8 kDa) was subjected to gel filtration.…”

Section: Resultsmentioning

confidence: 99%

Enzymatic Properties of Futalosine Hydrolase, an Enzyme Essential to a Newly Identified Menaquinone Biosynthetic Pathway

Hiratsuka

Itoh

Seto

et al. 2009

Bioscience, Biotechnology, and Biochemistry

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“…In contrast, specific NH prefer purine nucleosides, being poor catalysts for pirimidine nucleosides and p-nitrophenyl--D-ribofuranoside hydrolysis. [29][30][31][32] The monomeric NH from L. donovani (LdNH) presents 314 residues and its three-dimensional structure has not been experimentally elucidated yet. Fortunately it presents a quite significant homology with the nonspecific Leishmania major nucleoside hydrolase (LmNH), 95% of sequential identity, and Crithidia fasciculata nucleoside hydrolase (CfNH), 80% of sequential identity, suggesting that this enzyme is also nonspecific, as proposed by Cui et al 16 based on enzymatic assays.The high conservation of the NH sequences among the Trypanosomatidae species suggests that this enzyme is fundamental for the purines and pirimidines salvage in these species.…”

mentioning

confidence: 99%

Design of inhibitors for nucleoside hydrolase from Leishmania donovani using molecular dynamics studies

França¹,

Rocha²,

Reboredo³

et al. 2008

J. Braz. Chem. Soc.

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Neste trabalho é proposto o primeiro modelo por homologia para a nucleosídeo hidrolase de Leishmania donovani construído a partir das estruturas das nucleosídeo hidrolases de Crithidia fasciculata e de Leishmania major. Usando as informações de interação entre o inibidor p-aminofeniliminoribitol e a nucleosídeo hidrolase de Crithidia fasciculata foram planejados dois novos potenciais inibidores, os quais apresentam novas interações com alguns resíduos da bolsa hidrofóbica do sítio ativo do modelo. Simulações por dinâmica molecular dos protótipos ancorados nos sítios ativos do modelo e das enzimas usadas como moldes, mostraram que, diferente do p-aminofeniliminoribitol, eles permaneceram ancorados nos sítios ativos das três enzimas ao longo de toda a dinâmica, interagindo fortemente com os aminoácidos da bolsa hidrofóbica.In this work we propose the first homology model for nucleoside hydrolase from Leishmania donovani, built based on the crystallographic structures of Crithidia fasciculata and Leishmania major nucleoside hydrolases. We used the interaction information from the crystallographic model of the enzyme of C. fasciculata in complex with the inhibitor p-aminophenyliminoribitol, to design two new potential inhibitors, which present new interactions with some residues of the hydrophobic pocket of the model active site. Molecular dynamics simulations of the prototypes inside the active sites of the model and the template enzymes showed that, differently from p-aminophenyliminoribitol, they remained tightly bound inside the active sites, interacting strongly with the amino acids from the hydrophobic pocket. Keywords: visceral leishmaniasis, Leishmania donovani, metalloproteins, homology modeling, molecular dynamics IntroductionAccording to the World Health Organization (WHO), leishmaniasis is a threat for 350 million people in 88 countries, where 72 of those are developing countries. WHO also estimates that 12 million people in the world are infected and 2 million new cases appear annually, despite the official report of only about 600,000 new cases every year. 1 It is clear that leishmaniasis is one of the most important parasitic diseases, which is caused by protozoan of the Leishmania genus. This disease affects humans in four different forms with a broad range of clinical manifestations: visceral, mucocutaneous, cutaneous and diffuse cutaneous leishmaniasis. [1][2][3][4] Visceral leishmaniasis, also known as "kala-azar", is considered the most dangerous form of this disease, being fatal in 100% of the untreated cases 1 and is caused by the L. donovani complex. 4The search for prevention against leishmaniasis infection includes vaccines with weakened and genetically modified strains of the parasite and recombinant antigens 5 or its encoding DNA. 6 The only preventive treatment for leishmaniasis available today is the canine vaccine 65 França et al. Vol. 19, No. 1, 2008 Leishmune ® . 7 An efficient human vaccine is not available yet. Accordingly, the most effective way to combat leishmaniasis stil...

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Purine-specific Nucleoside N-Ribohydrolase from Trypanosoma brucei brucei

Cited by 90 publications

References 25 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

Enzymatic Properties of Futalosine Hydrolase, an Enzyme Essential to a Newly Identified Menaquinone Biosynthetic Pathway

Design of inhibitors for nucleoside hydrolase from Leishmania donovani using molecular dynamics studies

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