scholarly journals IDH1 and IDH2 mutants identified in cancer lose inhibition by isocitrate because of a change in their binding sites

2018 ◽  
Author(s):  
Juan P. Bascur ◽  
Melissa Alegría-Arcos ◽  
Ingrid Araya-Durán ◽  
Ezequiel I. Juritz ◽  
Fernando D. González-Nilo ◽  
...  
Keyword(s):  
Molecular Docking ◽  
Binding Sites ◽  
Competitive Inhibition ◽  
Binding Energies ◽  
Substrate Affinity ◽  
Wild Type ◽  
Similarity Estimation ◽  
In The Wild ◽  
Docking Calculations ◽  
Idh1 And Idh2 Mutations

AbstractIDH1 and IDH2 are human enzymes that convert isocitrate (ICT) into α-ketoglutarate (AKG). However, mutations in positions R132 of IDH1 and R140 and R172 of IDH2 cause these enzymes to convert AKG into 2-hydroxyglutarate (2HG). Concurrently, accumulation of 2HG in the cell is correlated with the development of cancer. This activity change is mainly due to the loss of the competitive inhibition by ICT of these enzymes, but the molecular mechanism behind this loss of inhibition is currently unknown. In this work we characterized the inhibition and loss of inhibition of IDH1 and IDH2 by means of the binding energies derived from molecular docking calculations. We characterized the substrate binding sites and how they differ among the mutant and wild type enzymes using a Jaccard similarity coefficient based on the residues involved in binding the substrates. We found that molecular docking effectively identifies the inhibition by ICT in the wild type and mutant enzymes that do not appear in tumors, and the loss of inhibition in the mutant enzymes that appear in tumors. Additionally, we found that the binding sites of the mutant enzymes are different among themselves. Finally, we found that the regulatory segment of IDH1 plays a prominent role in the change of binding sites between the mutant enzymes and the wild-type enzymes. Our findings show that the loss of inhibition is related to variations in the enzyme binding sites. Additionally, our findings show that a drug capable of targeting all IDH1 and IDH2 mutations in cancer is unlikely to be found due to significant differences among the binding sites of these paralogs. Moreover, the methodology developed here, which combines molecular docking calculations with binding site similarity estimation, can be useful for engineering enzymes, for instance, when aiming to modify the substrate affinity of an enzyme.

scholarly journals Mutation Spectrum in TPO Gene of Bangladeshi Patients with Thyroid Dyshormonogenesis and Analysis of the Effects of Different Mutations on the Structural Features and Functions of TPO Protein through In Silico Approach

2019 ◽  
Vol 2019 ◽  
pp. 1-18 ◽  
Author(s):  
Mst. Noorjahan Begum ◽  
Md Tarikul Islam ◽  
Shekh Rezwan Hossain ◽  
Golam Sarower Bhuyan ◽  
Mohammad A. Halim ◽  
...  
Keyword(s):  
Molecular Docking ◽  
In Silico ◽  
Homology Modelling ◽  
Structural Features ◽  
Binding Energies ◽  
Thyroid Peroxidase ◽  
Wild Type ◽  
Protein Activity ◽  
Prosthetic Group ◽  
Thyroid Dyshormonogenesis

Although thyroid dyshormonogenesis (TDH) accounts for 10-20% of congenital hypothyroidism (CH), the molecular etiology of TDH is unknown in Bangladesh. Thyroid peroxidase (TPO) is most frequently associated with TDH and the present study investigated the spectrum of TPO mutations in Bangladeshi patients and analyzed the effects of mutations on TPO protein structure through in silico approach. Sequencing-based analysis of TPO gene revealed four mutations in 36 diagnosed patients with TDH including three nonsynonymous mutations, namely, p.Ala373Ser, p.Ser398Thr, and p.Thr725Pro, and one synonymous mutation p.Pro715Pro. Homology modelling-based analysis of predicted structures of MPO-like domain (TPO142-738) and the full-length TPO protein (TPO1-933) revealed differences between mutant and wild type structures. Molecular docking studies were performed between predicted structures and heme. TPO1-933 predicted structure showed more reliable results in terms of interactions with the heme prosthetic group as the binding energies were -11.5 kcal/mol, -3.2 kcal/mol, -11.5 kcal/mol, and -7.9 kcal/mol for WT, p.Ala373Ser, p.Ser398Thr, and p.Thr725Pro, respectively, implying that p.Ala373Ser and p.Thr725Pro mutations were more damaging than p.Ser398Thr. However, for the TPO142-738 predicted structures, the binding energies were -11.9 kcal/mol, -10.8 kcal/mol, -2.5 kcal/mol, and -5.3 kcal/mol for the wild type protein, mutant proteins with p.Ala373Ser, p.Ser398Thr, and p.Thr725Pro substitutions, respectively. However, when the interactions between the crucial residues including residues His239, Arg396, Glu399, and His494 of TPO protein and heme were taken into consideration using both TPO1-933 and TPO142-738 predicted structures, it appeared that p.Ala373Ser and p.Thr725Pro could affect the interactions more severely than the p.Ser398Thr. Validation of the molecular docking results was performed by computer simulation in terms of quantum mechanics/molecular mechanics (QM/MM) and molecular dynamics (MD) simulation. In conclusion, the substitutions mutations, namely, p.Ala373Ser, p.Ser398Thr, and p.Thr725Pro, had been involved in Bangladeshi patients with TDH and molecular docking-based study revealed that these mutations had damaging effect on the TPO protein activity.

scholarly journals Analysis of a Dictyostelium chemotaxis mutant with altered chemoattractant binding

1988 ◽  
Vol 91 (4) ◽  
pp. 479-489 ◽  
Author(s):  
J.E. Segall ◽  
A.A. Bominaar ◽  
E. Wallraff ◽  
R.J. De Wit
Keyword(s):  
Linkage Group ◽  
Cyclic Amp ◽  
Cyclic Gmp ◽  
Binding Sites ◽  
Group Iv ◽  
Cell Area ◽  
Wild Type ◽  
High Affinity ◽  
In The Wild ◽  
Gmp Production

A Dictyostelium discoideum mutant defective in folate chemotaxis has been analysed using biochemical, behavioural, and genetic methods. A subset of the cell-surface folate binding sites appeared to be locked in a high-affinity state from which folate dissociated extremely slowly. Changes in cell area and motility induced by step increases in folate required 10- to 100-fold higher concentrations than in the wild type. Folate-stimulated cyclic GMP production was also altered. Chemotactic responses to cyclic AMP as well as cyclic AMP-stimulated cyclic GMP production were normal. The mutation responsible for the chemotaxis defect, termed folA1000, was localized to linkage group IV. The alterations in folate binding and sensitivity to folate co-localized with the folA1000 mutation. We conclude that the folA1000 mutation arrests the folate chemotaxis receptor in a high affinity state that can only poorly transduce folate binding into chemotactic responses.

scholarly journals Nuclear Export of mRNA by TAP/NXF1 Requires Two Nucleoporin-Binding Sites but Not p15

2002 ◽  
Vol 22 (15) ◽  
pp. 5405-5418 ◽  
Author(s):  
Isabelle C. Braun ◽  
Andrea Herold ◽  
Michaela Rode ◽  
Elisa Izaurralde
Keyword(s):  
Binding Sites ◽  
Nuclear Export ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Wild Type ◽  
Structural Domains ◽  
Export Activity ◽  
In The Wild ◽  
Pore Complexes ◽  
Directional Transport

ABSTRACT Metazoan NXF1/p15 heterodimers promote export of bulk mRNA through nuclear pore complexes (NPC). NXF1 interacts with the NPC via two distinct structural domains, the UBA-like domain and the NTF2-like scaffold, which results from the heterodimerization of the NTF2-like domain of NXF1 with p15. Both domains feature a single nucleoporin-binding site, and they act synergistically to promote NPC translocation. Whether the NTF2-like scaffold (and thereby p15) contributes only to NXF1/NPC association or is also required for other functions, e.g., to impart directionality to the export process by regulating NXF1/NPC or NXF1/cargo interactions, remains unresolved. Here we show that a minimum of two nucleoporin-binding sites is required for NXF1-mediated export of cellular mRNA. These binding sites can be provided by an NTF2-like scaffold followed by a UBA-like domain (as in the wild-type protein) or by two NTF2-like scaffolds or two UBA-like domains in tandem. In the latter case, the export activity of NXF1 is independent of p15. Thus, as for the UBA-like domain, the function of the NTF2-like scaffold is confined to nucleoporin binding. More importantly, two copies of either of these domains are sufficient to promote directional transport of mRNA cargoes across the NPC.

Molecular Docking Studies of Methamphetamine and Amphetamine- Related Derivatives as an Inhibitor against Dopamine Receptor

2020 ◽  
Vol 16 (2) ◽  
pp. 122-133
Author(s):  
Kobra Foroughi ◽  
Mehdi Khaksari ◽  
Asghar Shayannia
Keyword(s):  
Molecular Docking ◽  
Dopamine Receptor ◽  
Competitive Inhibition ◽  
Neurodegenerative Disorder ◽  
Inhibitory Effect ◽  
Docking Studies ◽  
Binding Energies ◽  
Dopamine Neurons ◽  
Degenerative Disorders ◽  
Pubchem Database

Background: The catecholamines such as dopamine, norepinephrine, and epinephrine are neurotransmitters that regulate different physiological functions of the central nervous system. Some evidence suggests that the degeneration of dopamine neurons in the substantia nigra contributes to Parkinson’s Disease (PD), which is a neurodegenerative disorder and it is responsible for the major symptoms of PD. It is suggested that replenishment of striatal dopamine through the oral administration of the dopamine precursor, levodopa, can compensate for the lack of endogenously produced dopamine. Some studies have shown competitive inhibition of dopamine receptor such as methamphetamine, and other amphetamine-related derivatives, which block dopamine receptor activity to uptake dopamine. Methods: In this study, 3D structures of amphetamine, methamphetamine, cocaine, methylphenidate, cathinone, MDMA, and mephedrone were obtained from the PubChem database, which has reported some evidence about their inhibitory effect with dopamine receptor. Then, these structures were provided for molecular docking analysis by Autodock Vina software. Eventually, the binding energies between docked dopamine receptor and them were calculated and their interactions were prognosticated. Results: Our results indicated that all chemicals can interact with dopamine receptor molecule in the active site of dopamine and the minimum binding energies belong to Cocaine and Methylphenidate with -7.9 Kcal/mol and -7.2 Kcal/mol, respectively. Conclusion: It might be concluded that amphetamine, methamphetamine, cocaine, methylphenidate, cathinone, MDMA, and mephedrone could act as potential inhibitors of DA receptor for dopamine uptake, which could cause degenerative disorders.

scholarly journals Atazanavir is a competitive inhibitor of SARS-CoV-2 Mpro, impairing variants replication in vitro and in vivo

2021 ◽  
Author(s):  
Otavio Augusto Chaves ◽  
Carolina Q. Sacramento ◽  
Andre Costa Ferreira ◽  
Mayara Mattos ◽  
Natalia Fintelman-Rodrigues ◽  
...  
Keyword(s):  
Molecular Docking ◽  
Competitive Inhibition ◽  
Competitive Inhibitor ◽  
Positive Control ◽  
Lethal Challenge ◽  
Angiotensin Converting Enzyme 2 ◽  
Main Protease ◽  
Docking Calculations

Atazanavir (ATV) has already been considered as a potential repurposing drug to 2019 coronavirus disease (COVID-19), however, there are controversial reports on its mechanism of action and effectiveness as anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Through the pre-clinical chain of experiments: enzymatic, molecular docking, cell-based, and in vivo assays, it is demonstrated here that both SARS-CoV-2 B.1 lineage and variant of concern gamma are susceptible to this antiretroviral. Enzymatic assays and molecular docking calculations showed that SARS-CoV-2 main protease (Mpro) was inhibited by ATV, with Morrison's inhibitory constant (Ki) 1.5-fold higher than boceprevir (GC376, a positive control). ATV was a competitive inhibition, increasing the Mpro's Michaelis-Menten (Km) more than 6-fold. Cell-based assays indicated that SARS-CoV-2 gamma is more susceptible to ATV than its predecessor strain B.1. Using oral administration of ATV in mice to reach plasmatic exposure similar to humans, transgenic mice expression in human angiotensin converting enzyme 2 (K18-hACE2) were partially protected against lethal challenge with SARS-CoV-2 gamma. Moreover, less cell death and inflammation were observed in the lung from infected and treated mice. Our studies may contribute to a better comprehension of the Mpro/ATV interaction, which could pave the way to the development of specific inhibitors of this viral protease.

Cleavage of Arpin by Calpain as a Potential Regulation Mechanism of the Arp2/3 Complex and Cell Motility

2018 ◽  
Author(s):  
Renee Potashner
Keyword(s):  
Plasma Membrane ◽  
Cell Motility ◽  
Cancer Metastasis ◽  
Competitive Inhibition ◽  
Cell Movement ◽  
Personal Communication ◽  
Cysteine Proteases ◽  
Regulation Mechanism ◽  
Wild Type ◽  
In The Wild

Cell movement is mediated by cycles of actin polymerisation. A novel protein, Arpin, was discovered that has been suggested to decrease cell motility through competitive inhibition of WASP family proteins, the activators of the complex driving actin polymerisation. In preliminary studies, Arpin was found to inhibit the Arp2/3 complex (Gautreau and Blanchoin, personal communication). Arpin contains sequence homology to the Arp2/3-binding site of WASP proteins. Calpains, a family of intracellular calcium-dependent cysteine proteases, can be found near the plasma membrane and the concentration of calcium ion required for activation is decreased when calpain is bound to the plasma membrane in the presence of phospholipids (Leloup et al. 2010). The common localization of calpain and Arpin, along with the known contributions calpain has in regulating other cell motility proteins, makes calpain a likely candidate for Arpin regulation. By transfecting calpain wild-type (pz/pz), knockdown (p/p) and lentivirus rescue mouse embryonic fibroblasts with a plasmid containing the Arpin gene (c15orf38), I hope to show the presence of differential cleavage of Arpin by calpain in the wild-type cells compared to the calpain knock-down cells through Western blot analysis. Understanding how Arpin is regulated will provide a basis for further research in cell motility, which has contributions to cancer metastasis and other diseases.  

scholarly journals IN-SILICO ALANINE SCANNING ANALYSIS ON THE CATALYTIC RESIDUES OF A NOVEL Β-GLUCOSIDASE FROM TRICHODERMA ASPERELLUM UC1

2021 ◽  
Vol 83 (3) ◽  
pp. 61-73
Author(s):  
Aimi Aliah Mohamad Yunus ◽  
Roswanira Abdul Wahab ◽  
Aina Hazimah Bahaman ◽  
Habeebat Adekilekun Oyewusi ◽  
Syariffah Nuratiqah Syed Yaacob
Keyword(s):  
Binding Energies ◽  
Preference Order ◽  
Trichoderma Asperellum ◽  
Wild Type ◽  
Intermolecular Hydrogen Bonds ◽  
Alanine Scanning ◽  
Scanning Method ◽  
Catalytic Residues ◽  
In The Wild ◽  
The Stability

Currently, the catalytic residue of the highly prolific fungal β-glucosidase (BGL) of Trichoderma asperellum UC1 remains unvalidated.  The study used the alanine scanning method to confirm the catalytic residues of the BGL as Glu165, Asp226, and Glu423. This method cancels out all intermolecular hydrogen bonds with substrates, lignin, hemicellulose, and cellulose. Results revealed an overall decline in the stability of the energy-minimized mutant enzymes' compared to the wild-type BGL. The mutant enzyme registered lower PROCHECK (91.0%), ERRAT (93.09%), and Verify-3D (98.92%) values, in comparison to 90.2%, 92.09%, 98.06%, in the wild-type BGL, respectively. The mutant BGL UC1-substrate complexes were less stable than the wild-type enzyme, in which the mutant exhibited higher binding energies for docked lignin (−7.4% kcal mol-1), cellulose (−7.2 kcal mol-1), and hemicellulose (−7.2 kcal mol-1). Binding energies of the wild-type BGL with the corresponding substrates were lower at −7.9 kcal mol-1, −8.1 kcal mol-1, and −7.8 kcal mol-1. An interesting observation was that the alanine scanning changed the substrate preference order based on the calculated binding energies. The mutant BGL bound preferentially to lignin>cellulose=hemicellulose, while the wild-type BGL was selective to cellulose>lignin>hemicellulose. Hence, the findings convey the high likelihood of Glu165, Asp 256, and Glu423 are the catalytic residues of the BGL of T. asperellum UC1.

scholarly journals Positive regulation of flhDC expression by OmpR in Yersinia pseudotuberculosis

Microbiology ◽  
2009 ◽  
Vol 155 (11) ◽  
pp. 3622-3631 ◽  
Author(s):  
Yangbo Hu ◽  
Yao Wang ◽  
Lisha Ding ◽  
Pei Lu ◽  
Steve Atkinson ◽  
...  
Keyword(s):  
Binding Sites ◽  
Yersinia Pseudotuberculosis ◽  
Bacterial Species ◽  
Bacterial Motility ◽  
Wild Type ◽  
Mobility Shift ◽  
Positive Regulation ◽  
In The Wild ◽  
Mobility Shift Assay ◽  
Extension Analysis

OmpR has been demonstrated to negatively regulate the expression of the flagellar master operon flhDC in a wide variety of bacterial species. Here we report the positive regulation of flhDC expression by OmpR in Yersinia pseudotuberculosis. A σ 70-dependent promoter was identified by primer extension analysis and an active region with two conserved OmpR-binding sites around the flhDC promoter was confirmed. To confirm the regulation of flhDC expression by OmpR, flhDC as well as the downstream flagellar genes fliA, flgD, flgA, flgM, fliC and flaA were fused to lacZ, and decreased expression of all these genes in an ompR mutant (ΔompR) was detected. Furthermore, ΔompR was defective in bacterial motility and flagella synthesis. This defect was due to the low level of expression of flhDC in ΔompR since overproduction of FlhDC in ΔompR restored bacterial motility. The importance of two conserved OmpR-binding sites around the flhDC promoter region in the regulation of flhDC expression by OmpR was demonstrated by the fact that mutation of either one or both sites significantly decreased the promoter activity in the wild-type but not in ΔompR. The binding of OmpR to these two sites was also demonstrated by DNA mobility shift assay. The possible mechanism underlying this positive regulation in Y. pseudotuberculosis is discussed. To our knowledge, this is the first report to demonstrate that OmpR positively regulates flhDC expression.

scholarly journals Substrate Affinity and Specificity of the ScSth1p Bromodomain Are Fine-Tuned for Versatile Histone Recognition

2019 ◽  
Author(s):  
Bartlomiej J. Blus ◽  
Hideharu Hashimoto ◽  
Hyuk-Soo Seo ◽  
Aleksandra Krolak ◽  
Erik W. Debler
Keyword(s):  
Binding Sites ◽  
Biological Function ◽  
Point Mutations ◽  
Substrate Affinity ◽  
List Type ◽  
Wild Type ◽  
Wide Range ◽  
Lysine Residues ◽  
Loop Regions ◽  
Peptide Binding Affinity

SummaryBromodomains recognize a wide range of acetylated lysine residues in histones and other nuclear proteins. Substrate specificity is critical for their biological function and arises from unique acetyl-lysine binding sites formed by variable loop regions. Here, we analyzed substrate affinity and specificity of the yeast ScSth1p bromodomain, an essential component of the “Remodels the Structure of Chromatin” complex, and found that the wild-type bromodomain preferentially recognizes H3K14ac and H4K20ac peptides. Mutagenesis studies—guided by our crystal structure determined at 2.7 Å resolution—revealed loop residues Ser1276 and Trp1338 as key determinants for such interactions. Strikingly, point mutations of each of these residues substantially increased peptide binding affinity and selectivity, respectively. Our data demonstrate that the ScSth1p bromodomain is not optimized for binding to an individual acetylation mark, but fine-tuned for interactions with several such modifications, consistent with the versatile and multivalent nature of histone recognition by reader modules such as bromodomains.HighlightsThe ScSth1p bromodomain preferentially recognizes H3K14ac and H4K20ac peptidesSer1276 and Trp1338 are key determinants of substrate affinity and specificityMutations of these residues drastically increase substrate affinity and specificityThe ScSth1p bromodomain is fine-tuned for promiscuous histone tail recognitionGraphical Abstract

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