scholarly journals Differences in the subunit interface residues of alternatively spliced glutathione transferases affects catalytic and structural functions

2007 ◽  
Vol 401 (3) ◽  
pp. 635-644 ◽  
Author(s):  
Juthamart Piromjitpong ◽  
Jantana Wongsantichon ◽  
Albert J. Ketterman
Keyword(s):  
Active Site ◽  
Electrostatic Interactions ◽  
Hydrophobic Interactions ◽  
Glutathione Transferases ◽  
Active Site Residues ◽  
Interface Area ◽  
Subunit Interface ◽  
Alternatively Spliced ◽  
Active Site Conformation ◽  
Interface Residues

GSTs (glutathione transferases) are multifunctional widespread enzymes. Currently there are 13 identified classes within this family. Previously most structural characterization has been reported for mammalian Alpha, Mu and Pi class GSTs. In the present study we characterize two enzymes from the insect-specific Delta class, adGSTD3-3 and adGSTD4-4. These two proteins are alternatively spliced products from the same gene and have very similar tertiary structures. Several major contributions to the dimer interface area can be separated into three regions: conserved electrostatic interactions in region 1, hydrophobic interactions in region 2 and an ionic network in region 3. The four amino acid side chains studied in region 1 interact with each other as a planar rectangle. These interactions are highly conserved among the GST classes, Delta, Sigma and Theta. The hydrophobic residues in region 2 are not only subunit interface residues but also active site residues. Overall these three regions provide important contributions to stabilization and folding of the protein. In addition, decreases in yield as well as catalytic activity changes, suggest that the mutations in these regions can disrupt the active site conformation which decreases binding affinity, alters kinetic constants and alters substrate specificity. Several of these residues have only a slight effect on the initial folding of each subunit but have more influence on the dimerization process as well as impacting upon appropriate active site conformation. The results also suggest that even splicing products from the same gene may have specific features in the subunit interface area that would preclude heterodimerization.

scholarly journals Molecular docking and pharmacokinetic screening of eucalyptol (1,8 cineole) from eucalyptus essential oil against SARS-CoV-2

2020 ◽  
Vol 12 (3) ◽  
pp. 536-545
Author(s):  
Arun D. SHARMA ◽  
Inderjeet KAUR
Keyword(s):  
Molecular Docking ◽  
Essential Oil ◽  
Active Site ◽  
In Silico ◽  
Hydrophobic Interactions ◽  
Rna Virus ◽  
In Silico Analysis ◽  
Active Site Residues ◽  
Virus Family

SARS-CoV-2 (COVID-19), member of corona virus family, is a positive single stranded RNA virus. Due to lack of drugs it is spreading its tentacles across the world. Being associated with cough, fever, and respiratory distress, this disease caused more than 15% mortality worldwide. Mpro/3CLpro has recently been regarded as a suitable target for drug design due to its vital role in virus replication. The current study focused on the inhibitory activity of eucalyptol (1,8 cineole), an essential oil component from eucalyptus oil, against Mpro/3CLprofrom SARS-CoV-2. Till date there is no work is undertaken on in-silico analysis of this compound against Mpro/3CLproof SARS-CoV-2. Molecular docking studies were conducted by using 1-click dock tool and Patchdock analysis. In-silico absorption, distribution, metabolism, excretion and toxicity (ADMET) profile were also studied. The calculated parameters such as docking score indicated effective binding of eucalyptol to COVID-19 Mpro protein. Active site prediction revealed the involvement of active site residues in ligand binding. Interactions results indicated that, Mpro/3CLpro/eucalyptol complexes forms hydrophobic interactions. ADMET studies provided guidelines and mechanistic scope for identification of potent anti-COVID 19 drug. Therefore, eucalyptol may represent potential herbal treatment to act as COVID-19 Mpro/3CLproinhibitor, a finding which must be validated in vivo.

scholarly journals Activation and selectivity of OTUB-1 and OTUB-2 deubiquitinylases

2020 ◽  
Vol 295 (20) ◽  
pp. 6972-6982
Author(s):  
Dakshinamurthy Sivakumar ◽  
Vikash Kumar ◽  
Michael Naumann ◽  
Matthias Stein
Keyword(s):  
Active Site ◽  
Electrostatic Interactions ◽  
Active Form ◽  
Cysteine Proteases ◽  
Binding Pocket ◽  
Catalytic Triad ◽  
Catalytic Site ◽  
Dynamics Simulation ◽  
Active Site Residues ◽  
Catalytically Active

The ovarian tumor domain (OTU) deubiquitinylating cysteine proteases OTUB1 and OTUB2 (OTU ubiquitin aldehyde binding 1 and 2) are representative members of the OTU subfamily of deubiquitinylases. Deubiquitinylation critically regulates a multitude of important cellular processes, such as apoptosis, cell signaling, and growth. Moreover, elevated OTUB expression has been observed in various cancers, including glioma, endometrial cancer, ovarian cancer, and breast cancer. Here, using molecular dynamics simulation approaches, we found that both OTUB1 and OTUB2 display a catalytic triad characteristic of proteases but differ in their configuration and protonation states. The OTUB1 protein had a prearranged catalytic site, with strong electrostatic interactions between the active-site residues His265 and Asp267. In OTUB2, however, the arrangement of the catalytic triad was different. In the absence of ubiquitin, the neutral states of the catalytic-site residues in OTUB2 were more stable, resulting in larger distances between these residues. Only upon ubiquitin binding did the catalytic triad in OTUB2 rearrange and bring the active site into a catalytically feasible state. An analysis of water access channels revealed only a few diffusion trajectories for the catalytically active form of OTUB1, whereas in OTUB2 the catalytic site was solvent-accessible, and a larger number of water molecules reached and left the binding pocket. Interestingly, in OTUB2, the catalytic residues His224 and Asn226 formed a stable hydrogen bond. We propose that the observed differences in activation kinetics, protonation states, water channels, and active-site accessibility between OTUB1 and OTUB2 may be relevant for the selective design of OTU inhibitors.

Molecular docking and molecular dynamics simulations of fumarate hydratase and its mutant H235N complexed with pyromellitic acid and citrate

2017 ◽  
Vol 15 (06) ◽  
pp. 1750026 ◽  
Author(s):  
S. Subasri ◽  
Santosh Kumar Chaudhary ◽  
K. Sekar ◽  
Manish Kesherwani ◽  
D. Velmurugan
Keyword(s):  
Molecular Dynamics ◽  
Active Site ◽  
Model Building ◽  
Hydrophobic Interactions ◽  
Conformational Stability ◽  
Fumarate Hydratase ◽  
Md Simulations ◽  
Major Effect ◽  
Active Site Residues ◽  
Pyromellitic Acid

Fumarase catalyzes the reversible, stereospecific hydration/dehydration of fumarate to L-malate during the Kreb’s cycle. In the crystal structure of the tetrameric fumarase, it was found that some of the active site residues S145, T147, N188 G364 and H235 had water-mediated hydrogen bonding interactions with pyromellitic acid and citrate which help to the protonation state for the conversion of fumarate to malate. When His 235 is mutated with Asn (H235N), water-mediated interactions were lost due to the shifting of active site water molecule by 0.7 Å away. Molecular dynamics (MD) simulations were also carried out by NAMD and analyzed using Assisted Model Building with Energy Refinement (AMBER) program to better understand the conformational stability and other aspects during the binding of pyromellitic acid and citrate with native and mutant FH. The role of hydrogen bonds and hydrophobic interactions was also analyzed. The present study confirms that the H235N mutation has a major effect on the catalytic activity of fumarase which is evident from the biochemical studies.

scholarly journals Discovery of Fungal Metabolites Bergenin, Quercitrin and Dihydroartemisinin as Potential Inhibitors Against Main Protease of SARS-CoV-2

2020 ◽  
Author(s):  
Ravi Patel ◽  
Akash Vanzara ◽  
Nimisha Patel ◽  
Ajit Vasava ◽  
Sachin Patil ◽  
...  
Keyword(s):  
Bioactive Compounds ◽  
Active Site ◽  
Electrostatic Interactions ◽  
Hydrophobic Interactions ◽  
Binding Energies ◽  
Crystallographic Structure ◽  
Medicinal Uses ◽  
Main Protease ◽  
Cyathus Stercoreus ◽  
Potential Inhibitors

Emergence of severe acute respiratory syndrome corona virus 2 (SARS-CoV-2) infection has given rise to COVID-19 pandemic, that is wreaking havoc worldwide. Therefore, there is an urgent need to find out novel drugs to combat SARS-CoV-2 infection. In this backdrop, the present study was aimed to assess potent bioactive compounds from different fungi as potential inhibitors of SARS-CoV-2 main protease (M<sup>pro</sup>) using an <i>in-silico</i> analysis. Nearly 118 bioactive compounds were extracted from <i>Dictyophora indusiata</i>, <i>Geassstrum triplex</i> and <i>Cyathus stercoreus </i>and identified using HR LC/MS analysis. Of which, only bergenin (<i>D. indusiata</i>), quercitrin (<i>G. triplex</i>) and dihydroartemisinin (<i>C. stercoreus</i>) were selected based on their medicinal uses, binding score and active site covered. The 6LU7, a protein crystallographic structure of SARS-CoV-2 M<sup>pro</sup>, was docked with bergenin, quercitrin and dihydroartemisinin using Autodock 4.2 and the binding energies obtained were -7.86, -10.29 and -7.20 kcal/mol, respectively. Bergenin, quercitrin and dihydroartemisinin formed hydrogen bond, electrostatic interactions and hydrophobic interactions with foremost active site amino acids THR190, GLU166, GLN189, GLY143, HIS163, HIS164, CYS145 and PHE140. Present investigation suggests that these three drugs may be used as alternative inhibitors against SARS-CoV-2 M<sup>pro</sup>. However, further research is necessary to assess <i>in vitro</i> potential of these drugs. To the best of our knowledge, present investigation reported these three bioactive compounds of fungal origin for the first time.

Navigating Research Toward the Re-emerging Nipah Virus- A New Piece to the Puzzle

2019 ◽  
Vol 25 (12) ◽  
pp. 1392-1401
Author(s):  
Pritika Ramharack ◽  
Nikita Devnarain ◽  
Letitia Shunmugam ◽  
Mahmoud E.S. Soliman
Keyword(s):  
Active Site ◽  
Hydrophobic Interactions ◽  
Nipah Virus ◽  
Homology Model ◽  
Future Research ◽  
Active Site Residues ◽  
Comprehensive Overview ◽  
Computational Tools ◽  
Plot Analysis ◽  
Potential Inhibitors

Background: The recent Nipah virus (NiV) outbreak in India has caused a state of chaos, with potential to become the next international pandemic. There is still a great deal to learn about NiV for the development of a potent treatment against it. The NiV non-structural proteins play important roles in the lifecycle of the virus, with the RNA-dependent RNA-polymerase (RdRp) being a vital component in viral replication. In this study, we not only provide a comprehensive overview of all the literature concerning NiV, we also propose a model of the NiV RdRp and screen for potential inhibitors of the viral enzyme. Objectives: In this study, computational tools were utilized in the design of a NiV RdRp homology model. The active site of RdRp was then identified and potential inhibitors of the protein were discovered with the use of pharmacophore-based screening. Methods: In this study, computational tools were utilized in the design of a NiV RdRp homology model. The active site of RdRp was then identified and potential inhibitors of the protein were discovered with the use of pharmacophore-based screening. Results: Ramachandran plot analysis revealed a favourable model. Upon binding of nucleoside analog, 4’- Azidocytidine, active site residues Trp1714 and Ser1713 took part in stabilizing hydrogen bonds, while Thr1716, Ser1478, Ser1476 and Glu1465 contributed to hydrophobic interactions. Pharmacophore based screening yielded 18 hits, of which ZINC00085930 demonstrated the most optimal binding energy (-8.1 kcal/mol), validating its use for further analysis as an inhibitor of NiV. Conclusion: In this study we provide a critical guide, elucidating on the in silico requirements of the drug design and discovery process against NiV. This material lays a foundation for future research into the design and development of drugs that inhibit NiV.

scholarly journals Decoding the Molecular Effects of Atovaquone Linked Resistant Mutations on Plasmodium falciparum Cytb-ISP Complex in the Phospholipid Bilayer Membrane

2021 ◽  
Vol 22 (4) ◽  
pp. 2138
Author(s):  
Lorna Chebon-Bore ◽  
Taremekedzwa Allan Sanyanga ◽  
Colleen Varaidzo Manyumwa ◽  
Afrah Khairallah ◽  
Özlem Tastan Bishop
Keyword(s):  
Plasmodium Falciparum ◽  
Active Site ◽  
Protein Function ◽  
Hydrophobic Interactions ◽  
Point Mutations ◽  
Md Simulations ◽  
Resistance Mechanisms ◽  
Phospholipid Bilayer ◽  
Active Site Residues ◽  
Presence And Absence

Atovaquone (ATQ) is a drug used to prevent and treat malaria that functions by targeting the Plasmodium falciparum cytochrome b (PfCytb) protein. PfCytb catalyzes the transmembrane electron transfer (ET) pathway which maintains the mitochondrial membrane potential. The ubiquinol substrate binding site of the protein has heme bL, heme bH and iron-sulphur [2FE-2S] cluster cofactors that act as redox centers to aid in ET. Recent studies investigating ATQ resistance mechanisms have shown that point mutations of PfCytb confer resistance. Thus, understanding the resistance mechanisms at the molecular level via computational approaches incorporating phospholipid bilayer would help in the design of new efficacious drugs that are also capable of bypassing parasite resistance. With this knowledge gap, this article seeks to explore the effect of three drug resistant mutations Y268C, Y268N and Y268S on the PfCytb structure and function in the presence and absence of ATQ. To draw reliable conclusions, 350 ns all-atom membrane (POPC:POPE phospholipid bilayer) molecular dynamics (MD) simulations with derived metal parameters for the holo and ATQ-bound -proteins were performed. Thereafter, simulation outputs were analyzed using dynamic residue network (DRN) analysis. Across the triplicate MD runs, hydrophobic interactions, reported to be crucial in protein function were assessed. In both, the presence and absence of ATQ and a loss of key active site residue interactions were observed as a result of mutations. These active site residues included: Met 133, Trp136, Val140, Thr142, Ile258, Val259, Pro260 and Phe264. These changes to residue interactions are likely to destabilize the overall intra-protein residue communication network where the proteins’ function could be implicated. Protein dynamics of the ATQ-bound mutant complexes showed that they assumed a different pose to the wild-type, resulting in diminished residue interactions in the mutant proteins. In summary, this study presents insights on the possible effect of the mutations on ATQ drug activity causing resistance and describes accurate MD simulations in the presence of the lipid bilayer prior to conducting inhibitory drug discovery for the PfCytb-iron sulphur protein (Cytb-ISP) complex.

scholarly journals Discovery of Fungal Metabolites Bergenin, Quercitrin and Dihydroartemisinin as Potential Inhibitors Against Main Protease of SARS-CoV-2

2020 ◽  
Author(s):  
Ravi Patel ◽  
Akash Vanzara ◽  
Nimisha Patel ◽  
Ajit Vasava ◽  
Sachin Patil ◽  
...  
Keyword(s):  
Bioactive Compounds ◽  
Active Site ◽  
Electrostatic Interactions ◽  
Hydrophobic Interactions ◽  
Binding Energies ◽  
Crystallographic Structure ◽  
Medicinal Uses ◽  
Main Protease ◽  
Cyathus Stercoreus ◽  
Potential Inhibitors

Emergence of severe acute respiratory syndrome corona virus 2 (SARS-CoV-2) infection has given rise to COVID-19 pandemic, that is wreaking havoc worldwide. Therefore, there is an urgent need to find out novel drugs to combat SARS-CoV-2 infection. In this backdrop, the present study was aimed to assess potent bioactive compounds from different fungi as potential inhibitors of SARS-CoV-2 main protease (M<sup>pro</sup>) using an <i>in-silico</i> analysis. Nearly 118 bioactive compounds were extracted from <i>Dictyophora indusiata</i>, <i>Geassstrum triplex</i> and <i>Cyathus stercoreus </i>and identified using HR LC/MS analysis. Of which, only bergenin (<i>D. indusiata</i>), quercitrin (<i>G. triplex</i>) and dihydroartemisinin (<i>C. stercoreus</i>) were selected based on their medicinal uses, binding score and active site covered. The 6LU7, a protein crystallographic structure of SARS-CoV-2 M<sup>pro</sup>, was docked with bergenin, quercitrin and dihydroartemisinin using Autodock 4.2 and the binding energies obtained were -7.86, -10.29 and -7.20 kcal/mol, respectively. Bergenin, quercitrin and dihydroartemisinin formed hydrogen bond, electrostatic interactions and hydrophobic interactions with foremost active site amino acids THR190, GLU166, GLN189, GLY143, HIS163, HIS164, CYS145 and PHE140. Present investigation suggests that these three drugs may be used as alternative inhibitors against SARS-CoV-2 M<sup>pro</sup>. However, further research is necessary to assess <i>in vitro</i> potential of these drugs. To the best of our knowledge, present investigation reported these three bioactive compounds of fungal origin for the first time.

scholarly journals Electrostatic interactions affecting the active site of class Sigma glutathione S-transferase

2000 ◽  
Vol 347 (1) ◽  
pp. 193-197 ◽  
Author(s):  
Julie M. STEVENS ◽  
Richard N. ARMSTRONG ◽  
Heini W. DIRR
Keyword(s):  
Ionic Strength ◽  
Conformational Changes ◽  
Active Site ◽  
Electrostatic Interactions ◽  
Size Exclusion ◽  
Tryptophan Residue ◽  
Glutathione S Transferase ◽  
Protein Fluorescence ◽  
Subunit Interface ◽  
Α Helix

We have shown previously that the solvent-induced equilibrium unfolding mechanism of class Sigma glutathione S-transferase (GST) is strongly affected by ionic strength [Stevens, Hornby, Armstrong and Dirr (1998) Biochemistry 37, 15534-15541]. The protein is dimeric and has a hydrophilic subunit interface. Here we show that ionic strength alone has significant effects on the conformation of the protein, in particular at the active site. With the use of NaCl at up to 2 M under equilibrium conditions, the protein lost 60% of its catalytic activity and the single tryptophan residue per subunit became partly exposed. The effect was independent of protein concentration, eliminating the dissociation of the dimer as a possibility for the conformational changes. This was confirmed by size-exclusion HPLC. There was no significant change in the secondary structure of the protein according to far-UV CD data. Manual-mixing and stopped-flow kinetics experiments showed a slow single-exponential salt-induced change in protein fluorescence. For equilibrium and kinetics experiments, the addition of an active-site ligand (S-hexylglutathione) completely protected the protein from the ionic-strength-induced conformational changes. This suggests that the change occurs at or near the active site. Possible structural reasons for these novel effects are proposed, such as the flexibility of the α-helix 2 region as well as the hydrophilic subunit interface, highlighting the importance of electrostatic interactions in maintaining the structure of the active site of this GST.

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