Molecular basis of fosmidomycin's action on the human malaria parasite Plasmodium falciparum

Umeda, T.; Tanaka, Nobutada; Kusakabe, Yoshio; Nakanishi, Mahito; Kitade, Yukio; Nakamura, Kazuo

doi:10.1038/srep00009

Cited by 87 publications

(122 citation statements)

References 43 publications

(92 reference statements)

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“…Once a successful purification strategy had been developed, the importance of residues in the catalytic hatch and involved in cofactor binding could be studied using site-directed mutagenesis. The recent successful crystallization of PfDXR in the presence and absence of an inhibitor [24], coupled to the structural information that comes from rational protein engineering, will aid in the development of novel inhibitors. The PfDXR protein produced in this study will be used for ongoing research investigating novel inhibitors of this enzyme.…”

Section: Resultsmentioning

confidence: 99%

“…In the same year, the crystal structure of EcDXR in complex with NADPH was solved; and revealed the presence of a flexible loop that acted as a catalytic hatch that closed over the active site securing the substrate, and played an important role in the enzymatic reaction and substrate specificity [23]. Recently the crystal structure of PfDXR demonstrated an intrinsic flexibility of the enzyme to house inhibitors in the active site [24]. A model of PfDXR had been generated previously that showed structural similarity between E. coli and PfDXR, but the model was not publically available [25].…”

Section: Introductionmentioning

confidence: 99%

“…The three-dimensional structure of PfDXR in complex with an inhibitor and cofactor. Stick representation of a monomer of PfDXR with bound inhibitor FR900098 (an N-acetyl derivative of fosmidomycin) shown in green and NADPH in black, both denoted as stick models (pdb accession code: 3AUA [24]). Residues that are part of the catalytic hatch, K295 and K297 (coloured blue and represented as sticks), and residues involved in the binding of NADPH, V114 and N115 (coloured red and represented as sticks), were mutated to resemble EcDXR.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

The Druggable Antimalarial Target PfDXR: Overproduction Strategies and Kinetic Characterization

Goble¹,

Johnson²,

Ridder³

et al. 2013

Protein and Peptide Letters

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Plasmodium falciparum 1-deoxy-D-xylulose-5-phosphate reductoisomerase (PfDXR) is a key enzyme in the synthesis of isoprenoids in the malaria parasite, using a pathway that is absent in the human host. This enzyme is receiving attention as it has been validated as a promising drug target. However, an impediment to the characterisation of this enzyme has been the inability to obtain sufficient quantities of the enzyme in a soluble and functional form. The expression of PfDXR from the codon harmonised coding region, under conditions of strongly controlled transcription and induction, resulted in a yield of 2 -4 mg/L of enzyme, which is 8 to 10-fold higher than previously reported yields. The kinetic parameters K m , V max and k cat were determined for PfDXR using an NADPH-dependent assay. Residues K295 and K297, unique to species of Plasmodium and located in the catalytic hatch region; and residues V114 and N115, essential for NADPH binding, were mutated to resemble those found in E. coli DXR. Interestingly, these mutations decreased the substrate affinity of PfDXR to values resembling that of E. coli DXR. PfDXR-K295N, K297S and PfDXR-V114A, N115G demonstrated a decreased ability to turnover substrate by 4-fold and 2-fold respectively in comparison to PfDXR. This study indicates a difference in the role of the catalytic hatch in capturing substrate by species of Plasmodium. The results of this study could contribute to the development of inhibitors of PfDXR.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

The Druggable Antimalarial Target PfDXR: Overproduction Strategies and Kinetic Characterization

Goble¹,

Johnson²,

Ridder³

et al. 2013

Protein and Peptide Letters

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“…Hydrolysis of the tert-butyl ester group of 12a−c,f and 14d,e with 20% TFA in CH 2 Cl 2 and subsequent EDC- phosphonates 8a−d as before. 35 P-NMR spectra of 7a−f and 8a indicate that these appear as rotameric mixtures, a known 50,51 phenomenon that was further validated by variable-temperature 31 P-NMR studies.…”

Section: Synthesismentioning

confidence: 99%

Synthesis and Bioactivity of β-Substituted Fosmidomycin Analogues Targeting 1-Deoxy-d-xylulose-5-phosphate Reductoisomerase

et al. 2015

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ABSTRACT. In the wake of the escalating development of resistance to currently available drugs, blocking the vital MEP pathway for isoprenoid biosynthesis in pathogens such as Plasmodia or Mycobacteria offers interesting prospects for inhibiting their growth. Although the natural product retro-hydroxamate fosmidomycin and its homologue FR900098 potently inhibit 1-deoxy-D-xylulose-5-phosphate reductoisomerase (Dxr), a key enzyme in the MEP pathway, their in vivo activity is compromised due to poor absorption and a suboptimal pharmacokinetic profile. In an effort to facilitate cellular uptake, we introduced aryl or aralkyl substituents at the β-position of the hydroxamate analogue of FR900098. While direct addition of a β-aryl moiety resulted in poor inhibition, longer linkers between the carbon backbone and the phenyl ring were generally associated with better binding to the enzymes, as well as activity against P. falciparum.X-ray structures of the parasite Dxr-inhibitor complexes show that the modes of binding of the two classes of compounds are in fact different. When the "shorter" compounds were bound, the active site flap was fully ordered and similar to that observed for the retro-hydroxamates. The "longer" compounds generate a substantially different flap structure, in which a key tryptophan residue is displaced, and the aromatic group of the ligand lies between the tryptophan and the methyl group on the hydroxamate. Although the most promising new Dxr inhibitors lack activity against E. coli and M. smegmatis, they proved to be highly potent inhibitors of Plasmodium falciparum in vitro growth.3

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“…The crystal structure of PfDXR in complex with a fosmidomycin derivative was used for the generation of the complex-based pharmacophore model (Umeda et al 2011) using MOE (2014. The generated pharmacophore model was validated by a test database of 50 known inhibitors (Altincicek et al 2000;Singh et al 2007;Deng et al 2011;Jansson et al 2013) of PfDXR enzyme.…”

Section: Pharmacophore Model Generation and Validationmentioning

confidence: 99%

In silico identification of promiscuous scaffolds as potential inhibitors of 1-deoxy- d -xylulose 5-phosphate reductoisomerase for treatment of Falciparum malaria

Wadood

Ghufran

Hassan

et al. 2016

Pharmaceutical Biology

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Fosmidomycin is a promising DXR inhibitor, which showed safety as well as efficacy against Plasmodium falciparum malaria in clinical trials. However, due to its poor oral bioavailability and non-drug-like properties, the focus of medicinal chemists is to develop inhibitors with improved pharmacological properties. Objective: This study described the computational design of new and potent inhibitors for deoxyxylulose 5-phosphate reductoisomerase and the prediction of their pharmacokinetic and pharmacodynamic properties. Material and methods: A complex-based pharmacophore model was generated from the complex X-ray crystallographic structure of PfDXR using MOE (Molecular Operating Environment). Furthermore, MOE-Dock was used as docking software to predict the binding modes of hits and target enzyme.Results: Finally, 14 compounds were selected as new and potent inhibitors of PfDXR on the basis of pharmacophore mapping, docking score, binding energy and binding interactions with the active site residues of the target protein. The predicted pharmacokinetic properties showed improved permeability by efficiently crossing blood-brain barrier. While, in silico promiscuity binding data revealed that these hits also have the ability to bind with other P. falciparum drug targets. Discussion and conclusion: In conclusion, innovative scaffolds with novel modes of action, improved efficacy and acceptable physiochemical/pharmacokinetic properties were computationally identified.ARTICLE HISTORY

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Molecular basis of fosmidomycin's action on the human malaria parasite Plasmodium falciparum

Cited by 87 publications

References 43 publications

The Druggable Antimalarial Target PfDXR: Overproduction Strategies and Kinetic Characterization

The Druggable Antimalarial Target PfDXR: Overproduction Strategies and Kinetic Characterization

Synthesis and Bioactivity of β-Substituted Fosmidomycin Analogues Targeting 1-Deoxy-d-xylulose-5-phosphate Reductoisomerase

In silico identification of promiscuous scaffolds as potential inhibitors of 1-deoxy- d -xylulose 5-phosphate reductoisomerase for treatment of Falciparum malaria

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