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![Conserved functional residues of ADCs that bind to substrate. ( a ) Stereo view of structural superimposition of processed MtbADC (blue), processed Thermus thermophilus ADC complexed with substrate analog fumarate (red and PDB id: 2EEO) and Helicobacter pylori ADC complexed with substrate analog isoasparagine (green, PDB id: 1UHE). The conserved and interacting residues are labeled according to MtbADC and the interactions are shown as dashed lines. ( b ) Stereo view of the active site in the dimer interface. The figure was prepared using Molscript [36] and Raster3D [37]. doi:10.1371/journal.pone.0033521.g001](profile/Antonius-Vandongen/publication/223983039/figure/fig1/AS:305354412969984@1449813573506/Conserved-functional-residues-of-ADCs-that-bind-to-substrate-a-Stereo-view-of.png)
Conserved functional residues of ADCs that bind to substrate. ( a ) Stereo view of structural superimposition of processed MtbADC (blue), processed Thermus thermophilus ADC complexed with substrate analog fumarate (red and PDB id: 2EEO) and Helicobacter pylori ADC complexed with substrate analog isoasparagine (green, PDB id: 1UHE). The conserved and interacting residues are labeled according to MtbADC and the interactions are shown as dashed lines. ( b ) Stereo view of the active site in the dimer interface. The figure was prepared using Molscript [36] and Raster3D [37]. doi:10.1371/journal.pone.0033521.g001
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L-aspartate α-decarboxylase (ADC) belongs to a class of pyruvoyl dependent enzymes and catalyzes the conversion of aspartate to β-alanine in the pantothenate pathway, which is critical for the growth of several micro-organisms, including Mycobacterium tuberculosis (Mtb). Its presence only in micro-organisms, fungi and plants and its absence in anim...
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Citations
... The protein was prepared using the protein preparation wizard of Schrödinger. Default parameters were used for assigning bond orders, and the OPLS 2005 force field with an average RMSD value of 0.30 Å was used to optimize and minimize the crystal structure [21]. The receptor grid for 2IE0 was generated using receptor grid generation in Maestro [22]. ...
A virtual screening protocol combining a 3D-QSAR model with molecular docking procedures and physicochemical properties was used to find novel inhibitors of enoyl-acyl carrier protein (ACP) reductase against Mycobacterium tuberculosis. Initially, 32 isoniazid analogues were collected from literature and investigated against the tuberculosis target (PDB Id: 2IE0) using molecular docking. The docking studies were used to position the inhibitors into the active site of enoyl-ACP reductase to derive a receptor-based 3D-QSAR model. The 3D-QSAR model was built and shown to be statistically significant, with a high predictive ability for the training (R 2 = 0.97) and test (Q 2 = 0.73) sets. The analysis of the contour cubes derived from the 3D-QSAR model revealed the chemical features necessary for the inhibition of the enoyl-ACP reductase enzyme. The model was then used for virtual screening with the aim of identifying new inhibitors of enoyl-ACP reductase and predicting their potential activity. Based on the results of the above studies, 10 new molecules were proposed as enoyl-ACP reductase inhibitors with high binding affinity, activity prediction, and favorable ADME properties.
... The Arg54 residue along with the pyruvoyl group are key actors within the active site, which is found at the interface of adjacent subunits of the tetramer. Specifically, Arg54 aids in substrate recognition through formation of a salt bridge with Asp's b-carboxylate, while the pyruvoyl group forms a transient imine with its amino group to serve as an electron sink that facilitates its decarboxylation (Sharma et al., 2012a;Sharma et al., 2012b). ...
... Sharma and co-workers used a cheminformatics approach to identify potential inhibitors of MtPanD (Sharma et al., 2012b;Sharma et al., 2012a). Using crystal structures of PanD enzymes from Thermus thermophilus and Helicobacter pylori with substrates bound in the active site, they found that these overlapped with the active site of the processed MtPanD to within 1 Å. ...
... A more refined screen was performed using Glide's Extra Precision (XP) mode, resulting in 28 compounds being selected for pharmacokinetics analysis using Qikprop. From these, seven lead compounds were identified as the most suitable candidates for further testing and investigation (Sharma et al., 2012b;Sharma et al., 2012a). An additional seven inhibitors previously identified against EcPanD were also tested (Williamson and Brown, 1979;Sharma et al., 2012a). ...
Despite decades of dedicated research, there remains a dire need for new drugs against tuberculosis (TB). Current therapies are generations old and problematic. Resistance to these existing therapies results in an ever-increasing burden of patients with disease that is difficult or impossible to treat. Novel chemical entities with new mechanisms of action are therefore earnestly required. The biosynthesis of coenzyme A (CoA) has long been known to be essential in Mycobacterium tuberculosis (Mtb), the causative agent of TB. The pathway has been genetically validated by seminal studies in vitro and in vivo. In Mtb, the CoA biosynthetic pathway is comprised of nine enzymes: four to synthesize pantothenate (Pan) from L-aspartate and a-ketoisovalerate; five to synthesize CoA from Pan and pantetheine (PantSH). This review gathers literature reports on the structure/ mechanism, inhibitors, and vulnerability of each enzyme in the CoA pathway. In addition to traditional inhibition of a single enzyme, the CoA pathway offers an antimetabolite strategy as a promising alternative. In this review, we provide our assessment of what appear to be the best targets, and, thus, which CoA pathway enzymes present the best opportunities for antitubercular drug discovery moving forward.
... Extending our efforts to unravel the trends of diseases [29][30][31][32], we employed a novel network-based approach to establish disease-disease relationships. In this approach, Covid-19 associated human target coding genes (TC-genes) are extracted and classi ed based on the pathways shared by them. ...
Millions of people have been forfeiting their lives due to SARS-CoV-2 infection. Most of them are patients suffering from comorbid complications. However, what makes these patients susceptible to mortality is unknown. For this, we employed a novel network-based approach to Covid-19 associated human target coding genes (TC-genes) overlapping with high relevant diseases to reveal the disease-disease relation. Classification of TC-genes in our study suggests that most of them participate in signal transduction, immune and neuronal systems. The network-based approach provides an insight into the mechanism involving the cascade of the TC-genes action that may drastically increase the reactive oxygen species (ROS). An increase in ROS triggers high oxidative stress and inflammation in the body through the cytokines storm. The cytokines storm set the burden on the comorbid patient by weakening the system that may lead to mortality. Our work highlights the TC-genes that may link Covid-19 to certain diseases. Collectively, the study indicates that selective TC-genes can carry out an overlapping role in seemingly distinct mechanisms. Besides, many mechanisms could independently affect selective targets. Oxidative stress and inflammation are the common processes present in severe Covid-19 patients. The approach demonstrates the potential to elucidate disease-disease relationship that can be applied to other diseases.
... The importance of ring based patterns can be indicated by recent investigations. Determining the preference of ring based structural pattern can aid in molecular design to accomplish a particular function [17,[19][20][21][22][23][24]. For example; quinoline, a versatile heterocyclic fused ring scaffold is common in cancer drugs [19]. ...
Clustering brings molecules having similar patterns together and is governed mainly by the structural features (SFs). The challenge is to cluster in such a way that the minimum number of groups with significant molecules having similar prevalent patterns comes together with minimal human intervention. Determining an automatic and reliable approach to cluster molecules is crucial for clinical assessment of medical conditions. Hypertension is one of such health conditions and anti-hypertensives (AHs) are the approved drugs to treat it. Here, an attempt has been made to cluster the AHs to identify the prominent patterns within a group. Principal component analysis (PCA) and k-means are well established independent algorithms, however, in this work, clustering is proceeded by PCA and is followed by one-way analysis of similarities (ANOSIM). The additional step of statistical relevance brings novelty in the final selection of the cluster. The latter highlights the significant difference between the two or more groups to enhance the clustering based on the similarity of the features within a group. Clustering of the United States Food and Drug Administration agency approved anti-hypertensives into six groups show a success rate of 94.73%. Kruskal-Wallis test for k = 6 suggest that there is a significant difference between the sample medians. Analysis of the cluster identifies the prominent pattern within a group. The average specificity and sensitivity achieved for k = 6 are 98.0 ± 1.6% and 97.3 ± 4.5%, respectively. A brief overview of the structural or functional relevance of molecules overlapping in a PCA plot for k = 6 has been discussed. The study is likely to be useful for a preliminary assessment to identify prevalent trends in SFs of AHs.
... Taken together, this evidence, not only accentuate the importance of pantothenate biosynthesis pathways for virulence, but also for survival of many bacterial species. Therefore, the pantothenate biosynthesis pathway is considered as a promising candidate for the development of safe and broad specific antimicrobial agents [20][21][22]. ...
Pantothenate is the metabolic precursor of Coenzyme A, an indispensable cofactor for many fundamental cellular processes. In this study, we show that many bacterial species have acquired multiple copies of pantothenate biosynthesis pathway genes via horizontal and vertical gene transfer events. Some bacterial species were also found to lack panE and panD genes, and depended on alternative enzymes/metabolic sources for pantothenate production. To shed light on the factors responsible for such dynamic evolutionary selections, the structural and functional characteristics of P. aeruginosa ketopantoate reductase (KPR), an enzyme that catalyzes the rate-limiting step and also the most duplicated, was investigated. A comparative analysis of apo and NADP+ bound crystal structures of P. aeruginosa KPR with orthologs, revealed that the residues involved in the interaction with specific phosphate moiety of NADP+ are relatively less conserved, suggesting dynamic evolutionary trajectories in KPRs for redox cofactor selection. Our structural and biochemical data also show that the specific conformational changes mediated by NADPH binding facilitate the cooperative binding of ketopantoate. From drastically reduced catalytic activity for NADH catalyzed the reaction with significantly higher KM of ketopantoate, it appears that the binding of ketopantoate is allosterically regulated to confer redox cofactor specificity. Altogether, our results, in compliance with earlier studies, not only depict the role of lateral gene transfer events in many bacterial species for enhancing pantothenate production but also highlight the possible role of redox cofactor balance in the regulation of pantothenate biosynthesis pathways.
... It is found in plants, fungi and microorganisms, including Mycobacterium, but not in animals. It catalyzes the formation of β-alanine from L-aspartate in bacteria (Sharma et al., 2012). Pantothenate (vitamin B5) is synthesized from β-alanine, which is the precursor of 4′-phosphopantetheine and coenzyme A (CoA) which plays an essential role in the metabolism and biosynthesis of fatty acids, tricarboxylic acid cycle, and biosynthesis of polypeptides (Gopalan et al., 2006). ...
... Indeed, there is recent interest in developing new drugs targeting the pantothenate synthesis pathway in M. tuberculosis. [26][27][28] In summary, we found that panD mutations are closely associated with POA resistance in POA-resistant mutants lacking pncA and rpsA mutations. Inducible PanD overexpression caused significant resistance to POA in M. tuberculosis. ...
Pyrazinamide (PZA) is a frontline anti-tuberculosis drug that plays a crucial role in the treatment of both drug-susceptible and multidrug-resistant tuberculosis (MDR-TB). PZA is a prodrug that is converted to its active form, pyrazinoic acid (POA), by a nicotinamidase/pyrazinamidase encoded by the pncA gene, the mutation of which is the major cause of PZA resistance. Although RpsA (ribosomal protein S1, involved in trans-translation) has recently been shown to be a target of POA/PZA, whole-genome sequencing has identified mutations in the panD gene encoding aspartate decarboxylase in PZA-resistant strains lacking pncA and rpsA mutations. To gain more insight into a possible new target of PZA, we isolated 30 POA-resistant mutants lacking mutations in pncA and rpsA from M. tuberculosis in vitro, and whole-genome sequencing of 3 mutants identified various mutations in the panD gene. Additionally, sequencing analysis revealed that the remaining 27 POA-resistant mutants all harbored panD mutations affecting the C-terminus of the PanD protein, with PanD M117I being the most frequent mutation (24/30, 80%). Conditional overexpression of panD from M. tuberculosis, M. smegmatis or E. coli, or of M. tuberculosis mutant PanD M117I, all conferred resistance to POA and PZA in M. tuberculosis. β-alanine and pantothenate, which are downstream products of PanD, were found to antagonize the antituberculosis activity of POA. In addition, the activity of the M. tuberculosis PanD enzyme was inhibited by POA at therapeutically relevant concentrations in a concentration-dependent manner but was not inhibited by the prodrug PZA or the control compound nicotinamide. These findings suggest that PanD represents a new target of PZA/POA. These results have implications for a better understanding of this peculiar persister drug and for the design of new drugs targeting M. tuberculosis persisters for improved treatment.
... The previously reported and newly identified compounds that were tested in the present study for inhibitory effect against ADC are shown inFig. 1 . Oxaloacetate (K1), DL-threo-b-hydroxyaspartate (K2), L-glutamate (K3), L-cysteic acid (K4), succinate (K5), Lserine (K6), and D-serine (K7) had been tested previously, while Dtartrate (I1, ZINC00895296), L-tartrate (I2, ZINC00895301), 2,4- dihydroxypyrimidine-5-carboxylate (I3, ZINC00901606), D-tagatose (I4, ZINC03830878), (4S)-1,3-thiazolidin-3-ium-4-carboxyl- ate (I5, ZINC00967474), a-D-arabinopyranose (I6, ZINC03606295), and 1,2-dihydropyrazolo[3,4-d]pyrimidin-4-one (I7, ZINC05177572) were tested for the first time, stimulated by the hits recently identified in the chemoinformatics study [9]. The compounds were purchased from Sigma-Aldrich, Germany (K1– K7 and I1–I4), or from Labotest KG, Germany (I5–I7), in the highest commercially available purity (.97%). ...
... The inhibitory effects of these molecules have been previously quantified on E. coli ADC and inhibition constants have been determined in the mM range [10]. In our recent study [9], 336,761 molecules from three public databases have been targeted: the Maybridge (14,400 molecules), the Zinc (including the National Cancer Institute, 316,181 molecules), and the United States of America Foods and Drug Administration approved drugs database (3,180 molecules). A high-throughput virtual screening, based on the interaction of these compounds with the active site residues of the enzyme by using a stronger binding affinity as compared to that of the MtbADC:fumarate reference (24.2 kcal/mol) as a positive criterion, led to the identification of 28 lead molecules. ...
... Based on the crystal structure [6], the affinity of molecules to the active site of ADC was postulated to be dominated by two moieties: the pyruvoyl group and the Arg54 residue. All newly tested compounds were selected upon ensuring both a direct interaction with Arg54 residue, while excluding contact with the MtbADC pyruvoyl group [9]. Our data clearly reveal that the inhibitory potential of the tested ligands is, actually, directly dependent on the number of carboxylate groups present. ...
The catalytic activity of L-aspartate α-decarboxylase (ADC) is essential for the growth of several micro-organisms, including Mycobacterium tuberculosis (Mtb), and has triggered efforts for the development of pharmaceutically active compounds against tuberculosis. The present study is a continuation of our recent chemoinformatics-based design approach for identifying potential drug-like inhibitors against MtbADC. We report an NMR-based protocol that allows label-free and direct monitoring of enzymatic conversion, which we have combined with a systematic testing of reported and newly identified potential inhibitors against MtbADC. Quantification of enzymatic conversion in the absence and presence of inhibitors allowed for a relative measure of the inhibitory effect (k(rel)). Among the newly identified compounds, D-tartrate, L-tartrate, and 2,4-dihydroxypyrimidine-5-carboxylate were found to inhibit the enzyme with k(rel) values of 0.36, 0.38, and 0.54, respectively. In addition to the identification of potential building blocks for the development of therapeutic agents, the current study highlights the importance of electrostatic interactions governing enzyme-inhibitor binding.
Effective eradication therapy for Helicobacter pylori is a worldwide demand. Aspartate α-decarboxylase (ADC) was reported as a drug target in H. pylori, in an in silico study, with malonic acid (MA) as its inhibitor. We evaluated eradicating H. pylori infection through ADC inhibition and the possibility of resistance development. MA binding to ADC was modeled via molecular docking. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of MA were determined against H. pylori ATCC 43504, and a clinical H. pylori isolate. To confirm selective ADC inhibition, we redetermined the MIC in the presence of products of the inhibited enzymatic pathway: β-alanine and pantothenate. HPLC was used to assay the enzymatic activity of H. pylori 6x-his tagged ADC in the presence of different MA concentrations. H. pylori strains were serially exposed to MA for 14 passages, and the MICs were determined. Cytotoxicity in different cell lines was tested. The efficiency of ADC inhibition in treating H. pylori infections was evaluated using a Sprague–Dawley (SD) rat infection model. MA spectrum of activity was determined in different pathogens. MA binds to H. pylori ADC active site with a good docking score. The MIC of MA against H. pylori ranged from 0.5 to 0.75 mg/mL with MBC of 1.5 mg/mL. Increasing β-alanine and pantothenate concentrations proportionally increased MA MIC. The 6x-his tagged ADC activity decreased by increasing MA concentration. No resistance to ADC inhibition was recorded after 14 passages; MA lacked cytotoxicity in all tested cell lines. ADC inhibition effectively eradicated H. pylori infection in SD rats. MA had MIC between 0.625 to 1.25 mg/mL against the tested bacterial pathogens. In conclusion, ADC is a promising target for effectively eradicating H. pylori infection that is not affected by resistance development, besides being of broad-spectrum presence in different pathogens. MA provides a lead molecule for the development of an anti-helicobacter ADC inhibitor. This provides hope for saving the lives of those at high risk of infection with the carcinogenic H. pylori.
Background
α‐Amylase and α‐glucosidase are important therapeutic targets for the management of type 2 diabetes mellitus. The inhibition of these enzymes decreases postprandial hyperglycemia. In the present study compounds found in commercially available herbs and spices, were tested for their ability to inhibit α‐amylase and α‐glucosidase. These compounds were acetyleugenol, apigenin, cinnamic acid, eriodictyol, myrcene, piperine, and rosmarinic acid.
Methods
The enzyme inhibitory nature of the compounds was evaluated using in silico docking analysis with Maestro software and was further confirmed by in vitro α‐amylase and α‐glucosidase biochemical assays.
Results
The relationship between the in silico and in vitro results were well correlated, a more negative docking score was associated with a higher in vitro inhibitory activity. There was no significant (p > 0.05) difference between the Ki value of acarbose, a widely prescribed α‐glucosidase and α‐amylase inhibitor, and those of apigenin, eriodictyol, and piperine. For α‐amylase, there was no significant (p > 0.05) difference between the Ki value of acarbose and those of apigenin, cinnamic acid, and rosmarinic acid. The effect of the herbal compounds on cell viability was assessed with the SRB assay, in C2C12 and HepG2 cells. Acetyleugenol, cinnamic acid, myrcene, piperine, and rosmarinic acid had similar (p > 0.05) IC50 values to acarbose.
Conclusion
Several of the herbal compounds studied could regulate postprandial hyperglycemia. Using herbal plants has several advantages including low cost, natural origin and easy cultivation. These compounds can easily be consumed as teas or as herbs and spices to flavor food.
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