Using Fragment Cocktail Crystallography To Assist Inhibitor Design of <i>Trypanosoma brucei </i>Nucleoside 2-Deoxyribosyltransferase

Bosch, Jürgen; Robien, M.A.; Mehlin, Christopher; Boni, Erica; Riechers, Aaron; Buckner, Frederick S.; Voorhis, Wesley C. Van; Myler, Peter J.; Worthey, Elizabeth A.; DeTitta, George T.; Luft, Joseph R.; Lauricella, Angela; Gulde, Stacey; Anderson, L. L.; Kalyuzhniy, Oleksandr; Neely, H.; Ross, Jenni; Earnest, Thomas; Soltis, M.; Schoenfeld, Lori W.; Zucker, Frank; Merritt, E.A.; Fan, Erkang; Verlinde, Christophe L. M. J.; Hól, Wim G. J.

doi:10.1021/jm060429m

Cited by 67 publications

(62 citation statements)

References 30 publications

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“…19 It precisely defined the structure of the active site and the specific protein-nucleotide interactions. A similar catalytic triad composed of a tyrosine, an aspartate, and a glutamate is present in Ll NDT (Y7, D72, and E98) and in Rcl (Y13, D69, and E93) (Fig.…”

Section: Discussionmentioning

confidence: 99%

Structural Characterization of the Mammalian Deoxynucleotide N-Hydrolase Rcl and Its Stabilizing Interactions with Two Inhibitors

Yang

Padilla

Zhang

et al. 2009

Journal of Molecular Biology

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Section: Discussionmentioning

confidence: 99%

Structural Characterization of the Mammalian Deoxynucleotide N-Hydrolase Rcl and Its Stabilizing Interactions with Two Inhibitors

Yang

Padilla

Zhang

et al. 2009

Journal of Molecular Biology

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“…Such an approach has been reported in inhibitor design against the bacterial enzyme nucleoside 2-deoxyribosyltransferase. 13 …”

Section: Introductionmentioning

confidence: 99%

Small Molecule Binding Sites on the Ras:SOS Complex Can Be Exploited for Inhibition of Ras Activation

et al. 2015

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Constitutively active mutant KRas displays a reduced rate of GTP hydrolysis via both intrinsic and GTPaseactivating protein-catalysed mechanisms, resulting in the perpetual activation of Ras pathways. We describe a fragment screening campaign using X-ray crystallography that led to the discovery of three fragment binding sites on the Ras:SOS complex. The identification of tool compounds binding at each of these sites allowed exploration of two new approaches to Ras pathway inhibition by stabilising or covalently modifying the Ras:SOS complex to prevent the reloading of Ras with GTP. Initially, we identified ligands that bound reversibly to the Ras:SOS complex in two distinct sites, but these compounds were not sufficiently potent inhibitors to validate our stabilisation hypothesis. We conclude by demonstrating that covalent modification of Cys118 on Ras leads to a novel mechanism of inhibition of the SOS-mediated interaction between Ras and Raf, and is effective at inhibiting the exchange of labelled GDP in both mutant (G12C and G12V) and wild type Ras.

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“…ent in some species of Streptococcus (2) and in parasitic unicellular eukaryotic organisms such as Crithidia luciliae (3) and Trypanosoma brucei (4).…”

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confidence: 99%

In Vivo Reshaping the Catalytic Site of Nucleoside 2′-Deoxyribosyltransferase for Dideoxy- and Didehydronucleosides via a Single Amino Acid Substitution

Kaminski

Dacher

Dugué

et al. 2008

Journal of Biological Chemistry

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Nucleoside 2-deoxyribosyltransferases catalyze the transfer of 2-deoxyribose between bases and have been widely used as biocatalysts to synthesize a variety of nucleoside analogs. The genes encoding nucleoside 2-deoxyribosyltransferase (ndt) from Lactobacillus leichmannii and Lactobacillus fermentum underwent random mutagenesis to select variants specialized for the synthesis of 2,3-dideoxynucleosides. An Escherichia coli strain, auxotrophic for uracil and unable to use 2,3-dideoxyuridine, cytosine, and 2,3-dideoxycytidine as a source of uracil was constructed. Randomly mutated lactobacilli ndt libraries from two species, L. leichmannii and L. fermentum, were screened for the production of uracil with 2,3-dideoxyuridine as a source of uracil. Several mutants suitable for the synthesis of 2,3-dideoxynucleosides were isolated. The nucleotide sequence of the corresponding genes revealed a single mutation (G 3 A transition) leading to the substitution of a small aliphatic amino acid by a nucleophilic one, A15T (L. fermentum) or G9S (L. leichmannii), respectively. We concluded that the "adaptation" of the nucleoside 2-deoxyribosyltransferase activity to 2,3-dideoxyribosyl transfer requires an additional hydroxyl group on a key amino acid side chain of the protein to overcome the absence of such a group in the corresponding substrate. The evolved proteins also display significantly improved nucleoside 2,3-didehydro-2,3-dideoxyribosyltransferase activity.Nucleoside 2Ј-deoxyribosyltransferases (NDTs 2 ; EC 2.4.2.6) catalyze the transfer of the deoxyribosyl group (dR) from one base (X) to another base (Y), in the following reaction: dX ϩ Y % dY ϩX (where dX and dY represent the corresponding 2Ј-deoxyribonucleosides). These enzymes initially found in Lactobacillus participate in nucleoside salvage (1). NDTs are also present in some species of Streptococcus (2) and in parasitic unicellular eukaryotic organisms such as Crithidia luciliae (3) and Trypanosoma brucei (4).Two different enzymes have been described in Lactobacillus helveticus and Lactobacillus leichmannii; they are NDT I, specific for purines, and NDT II, which transfers dR between purine-purine, pyrimidine-pyrimidine, and purine-pyrimidine (5-7). The transferase reaction proceeds via a ping-pong bi-bi mechanism in which the first product is released leaving a covalent deoxyribosyl-enzyme intermediate before the second substrate combines with it (5, 8, 9, ). The stereospecificity of the reaction (only the ␤ anomer is formed) and the tolerance of NDT II for structural variations in the acceptor base have been exploited to synthesize various nucleoside analogs (10 -14).Enzymatic synthesis catalyzed by NDTs (10, 14) has several advantages over chemical methods. Indeed, chemical glycosylation increases the difficulty of obtaining the correct regioselectivity generating few secondary products (15). Furthermore, the instability of some intermediates under the conditions used for synthetic manipulations may be problematic, as demonstrated by the acid lability of purine ...

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Using Fragment Cocktail Crystallography To Assist Inhibitor Design of Trypanosoma brucei Nucleoside 2-Deoxyribosyltransferase

Cited by 67 publications

References 30 publications

Structural Characterization of the Mammalian Deoxynucleotide N-Hydrolase Rcl and Its Stabilizing Interactions with Two Inhibitors

Structural Characterization of the Mammalian Deoxynucleotide N-Hydrolase Rcl and Its Stabilizing Interactions with Two Inhibitors

Small Molecule Binding Sites on the Ras:SOS Complex Can Be Exploited for Inhibition of Ras Activation

In Vivo Reshaping the Catalytic Site of Nucleoside 2′-Deoxyribosyltransferase for Dideoxy- and Didehydronucleosides via a Single Amino Acid Substitution

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