Biochemical and structural insights into 6-phosphogluconate dehydrogenase from Leishmania donovani

Jakkula, Pranay; Narsimulu, Bandigi; Qureshi, Insaf Ahmed

doi:10.1007/s00253-021-11434-4

Cited by 20 publications

(6 citation statements)

References 57 publications

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“…To investigate the oligomerization behavior of Ld6PGD, we performed size exclusion chromatography (SEC) under different conditions. Under native conditions, the SEC profiles of Ld6PGD wt revealed one stable peak representing a molecular mass of 94-111 kDa (Figure 1a), which is equivalent to the dimeric form of the Ld6PGD wt and in accordance with previously published SEC profiles from Leishmania 6PGDs [24,47]. Varying Ld6PGD concentrations did not affect the elution profile (Figure S2).…”

Section: Resultssupporting

confidence: 88%

“…Based on these values, we calculated a catalytic efficiency of 1.5 ± 0.02 µM −1 •s −1 for 6PG and of 2.2 ± 0.2 µM −1 •s −1 for NADP + (Table 1). These values are very similar to other 6PGDs characterized so far [47,49,57,58] and nearly identical to 6PGDs of the related Trypanosoma species T. cruzi and T. brucei [59,60], with which Leishmania 6PGD shares a high sequence identity of approximately 70%.…”

Section: Production Oligomerization Behavior and Kinetic Characteriza...supporting

confidence: 85%

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Structure of Leishmania donovani 6-Phosphogluconate Dehydrogenase and Inhibition by Phosphine Gold(I) Complexes: A Potential Approach to Leishmaniasis Treatment

Berneburg,

Stumpf,

Velten

et al. 2023

IJMS

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As unicellular parasites are highly dependent on NADPH as a source for reducing equivalents, the main NADPH-producing enzymes glucose 6-phosphate dehydrogenase (G6PD) and 6-phosphogluconate dehydrogenase (6PGD) of the pentose phosphate pathway are considered promising antitrypanosomatid drug targets. Here we present the biochemical characterization and crystal structure of Leishmania donovani 6PGD (Ld6PGD) in complex with NADP(H). Most interestingly, a previously unknown conformation of NADPH is visible in this structure. In addition, we identified auranofin and other gold(I)-containing compounds as efficient Ld6PGD inhibitors, although it has so far been assumed that trypanothione reductase is the sole target of auranofin in Kinetoplastida. Interestingly, 6PGD from Plasmodium falciparum is also inhibited at lower micromolar concentrations, whereas human 6PGD is not. Mode-of-inhibition studies indicate that auranofin competes with 6PG for its binding site followed by a rapid irreversible inhibition. By analogy with other enzymes, this suggests that the gold moiety is responsible for the observed inhibition. Taken together, we identified gold(I)-containing compounds as an interesting class of inhibitors against 6PGDs from Leishmania and possibly from other protozoan parasites. Together with the three-dimensional crystal structure, this provides a valid basis for further drug discovery approaches.

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Section: Resultssupporting

confidence: 88%

Section: Production Oligomerization Behavior and Kinetic Characteriza...supporting

confidence: 85%

Structure of Leishmania donovani 6-Phosphogluconate Dehydrogenase and Inhibition by Phosphine Gold(I) Complexes: A Potential Approach to Leishmaniasis Treatment

Berneburg,

Stumpf,

Velten

et al. 2023

IJMS

View full text Add to dashboard Cite

show abstract

“…Subsequently, quenching experiments confirmed the tryptophan residues to be majorly buried inside the core of the enzyme, and a similar phenomenon has also been observed in the modeled Ld RPE structure. The ANS binding study of Ld RPE revealed aggregation at acidic pH in comparison to the protein at neutral pH, which was also reported in other leishmanial proteins such as PdxK and 6PGDH . The uncharged urea molecules have more tendency to bind the unfolded state of the protein as compared to the folded one; our study has also represented the two-state transition of Ld RPE with urea that gradually leads to unfolding.…”

Section: Discussionsupporting

confidence: 79%

“…The ANS binding study of LdRPE revealed aggregation at acidic pH in comparison to the protein at neutral pH, which was also reported in other leishmanial proteins such as PdxK 34 and 6PGDH. 35 The uncharged urea molecules have more tendency to bind the unfolded state of the protein as compared to the folded one; 36 our study has also represented the two-state transition of LdRPE with urea that gradually leads to unfolding. The three-dimensional structure of LdRPE represents all common topological features as observed in previously reported RPE crystal structures of P. falciparum 11 and H. sapiens.…”

Section: Discussionmentioning

confidence: 99%

Biophysical and Structural Characterization of Ribulose-5-phosphate Epimerase from Leishmania donovani

Narsimulu

Qureshi²,

Jakkula

et al. 2021

ACS Omega

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Pentose phosphate pathway (PPP) plays a crucial role in the maintenance of NADPH/NADP + homeostasis and provides protection against oxidative stress through detoxification of the reactive oxygen species. Ribulose-5-phosphate epimerase (RPE) participates in catalysis of the interconversion of ribulose-5-phosphate (Ru5P) to xylulose-5-phosphate (Xu5P) during PPP, however the structural attributes of this enzyme are still underexplored in many human pathogens including leishmanial parasites. The present study focuses upon cloning, purification and characterization of RPE of Leishmania donovani ( Ld RPE) using various biophysical and structural approaches. Sequence analysis has shown the presence of trypanosomatid-specific insertions at the N-terminus that are absent in humans and other eukaryotes. Gel filtration chromatography indicated recombinant Ld RPE to exist as a dimer in the solution. Circular dichroism studies revealed a higher alpha helical content at physiological pH and temperature that comparatively varies with changing these parameters. Additionally, intrinsic fluorescence and quenching studies of Ld RPE have depicted that tryptophan residues are mainly buried in the hydrophobic regions, and the recombinant enzyme is moderately tolerant to urea. Moreover, homology modeling was employed to generate the three-dimensional structure of Ld RPE followed by molecular docking with the substrate, product, and substrate analogues. The modeled structure of Ld RPE unravelled the presence of conserved active site residues as well as a single binding pocket for the substrate and product, while an in silico study suggested binding of substrate analogues into a similar pocket with more affinity than the substrate. Additionally, molecular dynamics simulation analysis has deciphered complexes of Ld RPE with most of the ligands exhibiting more stability than its apo form and lesser fluctuations in active site residues in the presence of ligands. Altogether, our study presents structural insights into leishmanial RPE that could provide the basis for its implication to develop potent antileishmanials.

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“…Therefore, the upregulated oxidative branch of the PPP probably supplies parasites with ribose-5-phosphate (R5P), which produces RNA and DNA, and NADPH for use as a cellular reductant in cellular metabolism. Recently, PPP has become a very attractive and viable metabolic pathway in drug target research of leishmaniasis, especially some enzymes that are involved in it [ 47 – 50 ], which is expected to lead to the development of novel drugs against VL.…”

Section: Discussionmentioning

confidence: 99%

Metabolomics analysis of visceral leishmaniasis based on urine of golden hamsters

Yuan,

Chen,

Zhao

et al. 2023

Parasites Vectors

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Background Leishmaniasis is one of the most neglected tropical diseases and is spread mainly in impoverished regions of the world. Although many studies have focused on the host’s response to Leishmania invasion, relatively less is known about the complex processes at the metabolic level, especially the metabolic alterations in the infected hosts. Methods In this study, we conducted metabolomics analysis on the urine of golden hamsters in the presence or absence of visceral leishmaniasis (VL) using the ultra-performance liquid chromatography (UPLC) system tandem high-resolution mass spectrometer (HRMS). The metabolic characteristics of urine samples, along with the histopathological change and the parasite burden of liver and spleen tissues, were detected at 4 and 12 weeks post infection (WPI), respectively. Results Amino acid metabolism was extensively affected at both stages of VL progression. Meanwhile, there were also distinct metabolic features at different stages. At 4 WPI, the significantly affected metabolic pathways involved alanine, aspartate and glutamate metabolism, the pentose phosphate pathway (PPP), histidine metabolism, tryptophan metabolism and tyrosine metabolism. At 12 WPI, the markedly enriched metabolic pathways were almost concentrated on amino acid metabolism, including tyrosine metabolism, taurine and hypotaurine metabolism and tryptophan metabolism. The dysregulated metabolites and metabolic pathways at 12 WPI were obviously less than those at 4 WPI. In addition, seven metabolites that were dysregulated at both stages through partial least squares-discriminant analysis (PLS-DA) and receiver-operating characteristic (ROC) tests were screened to be of diagnostic potential. The combination of these metabolites as a potential biomarker panel showed satisfactory performance in distinguishing infection groups from control groups as well as among different stages of infection. Conclusion Our findings could provide valuable information for further understanding of the host response to Leishmania infection from the aspect of the urine metabolome. The proposed urine biomarker panel could help in the development of a novel approach for the diagnosis and prognosis of VL. Graphical Abstract

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Biochemical and structural insights into 6-phosphogluconate dehydrogenase from Leishmania donovani

Cited by 20 publications

References 57 publications

Structure of Leishmania donovani 6-Phosphogluconate Dehydrogenase and Inhibition by Phosphine Gold(I) Complexes: A Potential Approach to Leishmaniasis Treatment

Structure of Leishmania donovani 6-Phosphogluconate Dehydrogenase and Inhibition by Phosphine Gold(I) Complexes: A Potential Approach to Leishmaniasis Treatment

Biophysical and Structural Characterization of Ribulose-5-phosphate Epimerase from Leishmania donovani

Metabolomics analysis of visceral leishmaniasis based on urine of golden hamsters

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