INSIGHTS INTO A 3D HOMOLOGY MODEL OF ARYLESTERASE: THE KEY RESIDUES UPON PROTEIN-LIGAND DOCKING AND MM-PBSA CALCULATIONS

2011 ◽  
Vol 10 (02) ◽  
pp. 165-177 ◽  
Author(s):  
WEIWEI HAN ◽  
YE WANG ◽  
QUAN LUO ◽  
YAN FENG ◽  
XIAODI NIU
Keyword(s):  
Computational Study ◽  
Fold Increase ◽  
Homology Model ◽  
Ligand Docking ◽  
Wild Type ◽  
Kinetic Experiment ◽  
Dynamics Calculation ◽  
Key Residues ◽  
Structural Insights ◽  
Vdw Interaction

Arylesterases (E.C. 3.1.1.2) play an important role in nature, which show high enantioselectivity toward chemically and pharmaceutically important compound in addition to being environmentally friendly. The docking results indicate that Tyr34, Leu98, Ile64, Met36, and Ile38 have important contributions to the substrate binding and the side chains of these residues can provide a rather vdW interaction with the substrate. The molecular dynamics calculation of the free energy by MM-PBSA method implies that the S31A and S31G mutant enzymes show more stability as compared with the wild type, which is in harmony with the kinetic experiment that the 2-fold and 2.5-fold increase in the Km for S31A and S31G enzymes. From these results, we can conjecture that mutations of Ser31 make the active site more spacious than that of arylerases', and it can lead to the enzyme active. The new structural insights obtained from this computational study are expected to stimulate further biochemical studies on the structures and mechanisms of other members of the SGNH-hydrolases.

Application of Molecular Docking for the Development of Improved HIV-1 Reverse Transcriptase Inhibitors

2020 ◽  
Vol 16 ◽  
Author(s):  
Arash Soltani ◽  
Seyed Isaac Hashemy ◽  
Farnaz Zahedi Avval ◽  
Houshang Rafatpanah ◽  
Seyed Abdolrahim Rezaee ◽  
...  
Keyword(s):  
Reverse Transcriptase ◽  
Binding Capacity ◽  
Binding Pocket ◽  
Ligand Docking ◽  
Binding Modes ◽  
Wild Type ◽  
Parent Molecule ◽  
Discovery Studio ◽  
Approved Drugs ◽  
Hiv 1

Introoduction: Inhibition of the reverse transcriptase (RT) enzyme of human immunodeficiency virus (HIV) by low molecular weight inhibitors is still an active area of research. Here, protein-ligand interactions and possible binding modes of novel compounds with the HIV-1 RT binding pocket (the wild-type as well as Y181C and K103N mutants) were obtained and discussed. Methods: A molecular fragment-based approach using FDA-approved drugs were followed to design novel chemical derivatives using delavirdine, efavirenz, etravirine and rilpivirine as the scaffolds. The drug-likeliness of the derivatives was evaluated using Swiss-ADME. Then the parent molecule and derivatives were docked into the binding pocket of related crystal structures (PDB ID: 4G1Q, 1IKW, 1KLM and 3MEC). Genetic Optimization for Ligand Docking (GOLD) Suite 5.2.2 software was used for docking and the results analyzed in the Discovery Studio Visualizer 4. A derivative was chosen for further analysis, if it passed drug-likeliness and the docked energy was more favorable than that of its parent molecule. Out of the fifty-seven derivatives, forty-eight failed in druglikeness screening by Swiss-ADME or in docking stage. Results: The final results showed that the selected compounds had higher predicted binding affinities than their parent scaffolds in both wild-type and the mutants. Binding energy improvement was higher for the structures designed based on second-generation NNRTIs (etravirine and rilpivirine) than the first-generation NNRTIs (delavirdine and efavirenz). For example, while the docked energy for rilpivirine was -51 KJ/mol, it was improved for its derivatives RPV01 and RPV15 up to -58.3 and -54.5 KJ/mol, respectively. Conclusion: In this study, we have identified and proposed some novel molecules with improved binding capacity for HIV RT using fragment-based approach.

scholarly journals Functional expression of a yeast mitochondrial intron-encoded protein requires RNA processing at a conserved dodecamer sequence at the 3' end of the gene.

1989 ◽  
Vol 9 (4) ◽  
pp. 1507-1512 ◽  
Author(s):  
H Zhu ◽  
H Conrad-Webb ◽  
X S Liao ◽  
P S Perlman ◽  
R A Butow
Keyword(s):  
Genetic Analysis ◽  
Rna Processing ◽  
Fold Increase ◽  
Functional Expression ◽  
Rrna Gene ◽  
Yeast Mitochondria ◽  
Wild Type ◽  
Site Specific ◽  
Var1 Gene ◽  
A Site

All mRNAs of yeast mitochondria are processed at their 3' ends within a conserved dodecamer sequence, 5'-AAUAAUAUUCUU-3'. A dominant nuclear suppressor, SUV3-I, was previously isolated because it suppresses a dodecamer deletion at the 3' end of the var1 gene. We have tested the effects of SUV3-1 on a mutant containing two adjacent transversions within a dodecamer at the 3' end of fit1, a gene located within the 1,143-base-pair intron of the 21S rRNA gene, whose product is a site-specific endonuclease required in crosses for the quantitative transmission of that intron to 21S alleles that lack it. The fit1 dodecamer mutations blocked both intron transmission and dodecamer cleavage, neither of which was suppressed by SUV3-1 when present in heterozygous or homozygous configurations. Unexpectedly, we found that SUV3-1 completely blocked cleavage of the wild-type fit1 dodecamer and, in SUV3-1 homozygous crosses, intron conversion. In addition, SUV3-1 resulted in at least a 40-fold increase in the amount of excised intron accumulated. Genetic analysis showed that these phenotypes resulted from the same mutation. We conclude that cleavage of a wild-type dodecamer sequence at the 3' end of the fit1 gene is essential for fit1 expression.

scholarly journals The TonB system in Aeromonas hydrophila NJ-35 is essential for MacA2B2 efflux pump-mediated macrolide resistance

2021 ◽  
Vol 52 (1) ◽  
Author(s):  
Yuhao Dong ◽  
Qing Li ◽  
Jinzhu Geng ◽  
Qing Cao ◽  
Dan Zhao ◽  
...  
Keyword(s):  
Aeromonas Hydrophila ◽  
Efflux Pump ◽  
Fold Increase ◽  
Activity Level ◽  
Macrolide Resistance ◽  
Wild Type ◽  
Iron Deprivation ◽  
Pump Activity ◽  
Tonb System ◽  
Increased Sensitivity

AbstractThe TonB system is generally considered as an energy transporting device for the absorption of nutrients. Our recent study showed that deletion of this system caused a significantly increased sensitivity of Aeromonas hydrophila to the macrolides erythromycin and roxithromycin, but had no effect on other classes of antibiotics. In this study, we found the sensitivity of ΔtonB123 to all macrolides tested revealed a 8- to 16-fold increase compared with the wild-type (WT) strain, but this increase was not related with iron deprivation caused by tonB123 deletion. Further study demonstrated that the deletion of tonB123 did not damage the integrity of the bacterial membrane but did hinder the function of macrolide efflux. Compared with the WT strain, deletion of macA2B2, one of two ATP-binding cassette (ABC) types of the macrolide efflux pump, enhanced the sensitivity to the same levels as those of ΔtonB123. Interestingly, the deletion of macA2B2 in the ΔtonB123 mutant did not cause further increase in sensitivity to macrolide resistance, indicating that the macrolide resistance afforded by the MacA2B2 pump was completely abrogated by tonB123 deletion. In addition, macA2B2 expression was not altered in the ΔtonB123 mutant, indicating that any influence of TonB on MacA2B2-mediated macrolide resistance was at the pump activity level. In conclusion, inactivation of the TonB system significantly compromises the resistance of A. hydrophila to macrolides, and the mechanism of action is related to the function of MacA2B2-mediated macrolide efflux.

scholarly journals Nonrecombinant meiosis I nondisjunction in Saccharomyces cerevisiae induced by tRNA ochre suppressors.

Genetics ◽  
1989 ◽  
Vol 123 (1) ◽  
pp. 81-95 ◽  
Author(s):  
E J Louis ◽  
J E Haber
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Chromosome ◽  
Stop Codon ◽  
Fold Increase ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Nonsense Mutations ◽  
Chromosome Iii ◽  
Meiosis I ◽  
Chromosome Nondisjunction

Abstract The presence of the tRNA ochre suppressors SUP11 and SUP5 is found to induce meiosis I nondisjunction in the yeast Saccharomyces cerevisiae. The induction increases with increasing dosage of the suppressor and decreases in the presence of an antisuppressor. The effect is independent of the chromosomal location of SUP11. Each of five different chromosomes monitored exhibited nondisjunction at frequencies of 0.1%-1.1% of random spores, which is a 16-160-fold increase over wild-type levels. Increased nondisjunction is reflected by a marked increase in tetrads with two and zero viable spores. In the case of chromosome III, for which a 50-cM map interval was monitored, the resulting disomes are all in the parental nonrecombinant configuration. Recombination along chromosome III appears normal both in meioses that have no nondisjunction and in meioses for which there was nondisjunction of another chromosome. We propose that a proportion of one or more proteins involved in chromosome pairing, recombination or segregation are aberrant due to translational read-through of the normal ochre stop codon. Hygromycin B, an antibiotic that can suppress nonsense mutations via translational read-through, also induces nonrecombinant meiosis I nondisjunction. Increases in mistranslation, therefore, increase the production of aneuploids during meiosis. There was no observable effect of SUP11 on mitotic chromosome nondisjunction; however some disomes caused SUP11 ade2-ochre strains to appear white or red, instead of pink.

scholarly journals HPF1 remodels the active site of PARP1 to enable the serine ADP-ribosylation of histones

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Fa-Hui Sun ◽  
Peng Zhao ◽  
Nan Zhang ◽  
Lu-Lu Kong ◽  
Catherine C. L. Wong ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Active Site ◽  
Negative Charge ◽  
Structural Data ◽  
Dna Breaks ◽  
Adp Ribosylation ◽  
Local Conformation ◽  
Key Residues ◽  
Structural Insights

AbstractUpon binding to DNA breaks, poly(ADP-ribose) polymerase 1 (PARP1) ADP-ribosylates itself and other factors to initiate DNA repair. Serine is the major residue for ADP-ribosylation upon DNA damage, which strictly depends on HPF1. Here, we report the crystal structures of human HPF1/PARP1-CAT ΔHD complex at 1.98 Å resolution, and mouse and human HPF1 at 1.71 Å and 1.57 Å resolution, respectively. Our structures and mutagenesis data confirm that the structural insights obtained in a recent HPF1/PARP2 study by Suskiewicz et al. apply to PARP1. Moreover, we quantitatively characterize the key residues necessary for HPF1/PARP1 binding. Our data show that through salt-bridging to Glu284/Asp286, Arg239 positions Glu284 to catalyze serine ADP-ribosylation, maintains the local conformation of HPF1 to limit PARP1 automodification, and facilitates HPF1/PARP1 binding by neutralizing the negative charge of Glu284. These findings, along with the high-resolution structural data, may facilitate drug discovery targeting PARP1.

scholarly journals Phosphoglycolate has profound metabolic effects but most likely no role in a metabolic DNA response in cancer cell lines

2019 ◽  
Vol 476 (4) ◽  
pp. 629-643 ◽  
Author(s):  
Isabelle Gerin ◽  
Marina Bury ◽  
Francesca Baldin ◽  
Julie Graff ◽  
Emile Van Schaftingen ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cell Lines ◽  
Oxidative Dna Damage ◽  
Fold Increase ◽  
Metabolic Effects ◽  
Phosphoglycerate Mutase ◽  
Wild Type ◽  
Metabolic Disturbances ◽  
Phosphoglycolate Phosphatase ◽  
Damage Repair

Abstract Repair of a certain type of oxidative DNA damage leads to the release of phosphoglycolate, which is an inhibitor of triose phosphate isomerase and is predicted to indirectly inhibit phosphoglycerate mutase activity. Thus, we hypothesized that phosphoglycolate might play a role in a metabolic DNA damage response. Here, we determined how phosphoglycolate is formed in cells, elucidated its effects on cellular metabolism and tested whether DNA damage repair might release sufficient phosphoglycolate to provoke metabolic effects. Phosphoglycolate concentrations were below 5 µM in wild-type U2OS and HCT116 cells and remained unchanged when we inactivated phosphoglycolate phosphatase (PGP), the enzyme that is believed to dephosphorylate phosphoglycolate. Treatment of PGP knockout cell lines with glycolate caused an up to 500-fold increase in phosphoglycolate concentrations, which resulted largely from a side activity of pyruvate kinase. This increase was much higher than in glycolate-treated wild-type cells and was accompanied by metabolite changes consistent with an inhibition of phosphoglycerate mutase, most likely due to the removal of the priming phosphorylation of this enzyme. Surprisingly, we found that phosphoglycolate also inhibits succinate dehydrogenase with a Ki value of <10 µM. Thus, phosphoglycolate can lead to profound metabolic disturbances. In contrast, phosphoglycolate concentrations were not significantly changed when we treated PGP knockout cells with Bleomycin or ionizing radiation, which are known to lead to the release of phosphoglycolate by causing DNA damage. Thus, phosphoglycolate concentrations due to DNA damage are too low to cause major metabolic changes in HCT116 and U2OS cells.

scholarly journals Engineering Desiccation Tolerance inEscherichia coli

2000 ◽  
Vol 66 (4) ◽  
pp. 1680-1684 ◽  
Author(s):  
Daniela Billi ◽  
Deborah J. Wright ◽  
Richard F. Helm ◽  
Todd Prickett ◽  
Malcolm Potts ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Phase Transition ◽  
Freeze Drying ◽  
Fold Increase ◽  
Inescherichia Coli ◽  
Vibration Frequencies ◽  
Wild Type ◽  
Air Drying ◽  
Phase Transition Temperatures ◽  
Cell Dehydration

ABSTRACT Recombinant sucrose-6-phosphate synthase (SpsA) was synthesized inEscherichia coli BL21DE3 by using the spsA gene of the cyanobacterium Synechocystis sp. strain PCC 6803. Transformants exhibited a 10,000-fold increase in survival compared to wild-type cells following either freeze-drying, air drying, or desiccation over phosphorus pentoxide. The phase transition temperatures and vibration frequencies (PO stretch) in phospholipids suggested that sucrose maintained membrane fluidity during cell dehydration.

scholarly journals OmpR and LeuO Positively Regulate the Salmonella enterica Serovar Typhi ompS2 Porin Gene

2004 ◽  
Vol 186 (10) ◽  
pp. 2909-2920 ◽  
Author(s):  
Marcos Fernández-Mora ◽  
José Luis Puente ◽  
Edmundo Calva
Keyword(s):  
Salmonella Enterica ◽  
Type Strain ◽  
Wild Type Strain ◽  
Phosphorylation Site ◽  
Regulatory Region ◽  
Random Mutagenesis ◽  
Fold Increase ◽  
Upstream Regulatory Region ◽  
Wild Type ◽  
Transposon Insertion

ABSTRACT The Salmonella enterica serovar Typhi ompS2 gene codes for a 362-amino-acid outer membrane protein that contains motifs common to the porin superfamily. It is expressed at very low levels compared to the major OmpC and OmpF porins, as observed for S. enterica serovar Typhi OmpS1, Escherichia coli OmpN, and Klebsiella pneumoniae OmpK37 quiescent porins. A region of 316 bp, between nucleotides −413 and −97 upstream of the transcriptional start point, is involved in negative regulation, as its removal resulted in a 10-fold increase in ompS2 expression in an S. enterica serovar Typhi wild-type strain. This enhancement in expression was not observed in isogenic mutant strains, which had specific deletions of the regulatory ompB (ompR envZ) operon. Furthermore, ompS2 expression was substantially reduced in the presence of the OmpR D55A mutant, altered in the major phosphorylation site. Upon random mutagenesis, a mutant where the transposon had inserted into the upstream regulatory region of the gene coding for the LeuO regulator, showed an increased level of ompS2 expression. Augmented expression of ompS2 was also obtained upon addition of cloned leuO to the wild-type strain, but not in an ompR isogenic derivative, consistent with the notion that the transposon insertion had increased the cellular levels of LeuO and with the observed dependence on OmpR. Moreover, LeuO and OmpR bound in close proximity, but independently, to the 5′ upstream regulatory region. Thus, the OmpR and LeuO regulators positively regulate ompS2.

Abstract 344: Endothelin-1 Overexpression Exaggerates Diabetes-induced Endothelial Dysfunction

Hypertension ◽  
2014 ◽  
Vol 64 (suppl_1) ◽  
Author(s):  
Noureddine IDRIS KHODJA ◽  
Muhammad Oneeb Rehman Mian ◽  
Tlili Barhoumi ◽  
Sofiane Ouerd ◽  
Jordan Gornitsky ◽  
...  
Keyword(s):  
Type 1 Diabetes ◽  
Endothelial Dysfunction ◽  
Vascular Complications ◽  
Fold Increase ◽  
Wild Type ◽  
No Donor ◽  
Meclofenamic Acid ◽  
Fibronectin Expression ◽  
Endothelium Dependent Relaxation

Objective: Vascular disease associated with endothelial dysfunction is a major cause of morbidity in patients with type-1 diabetes. Endothelin (ET)-1 plays a role in diabetes-induced vascular complications, since ET-1 type A receptor blockade reduces diabetes-induced vascular injury. However, whether ET-1 contributes to diabetes-induced endothelial dysfunction remains unproven. We hypothesized that vascular ET-1 overexpression will exaggerate diabetes-induced endothelial dysfunction. Methods: Diabetes was induced by streptozotocin treatment (STZ, 55 mg/kg/day, ip) for 5 days in 6-week-old male wild-type (WT) mice and in mice overexpressing ET-1 restricted to the endothelium (eET-1). Mice were studied 14 weeks later. Blood was collected to determine glucose. Mesenteric artery reactivity and remodeling were evaluated using pressurized myography and aortic fibronectin expression by immnunofluorescence. Results: STZ-induced diabetes was confirmed by a 3-fold increase in glycemia in WT and eET-1 ( P <0.001). Diabetes impaired endothelium-dependent relaxation (EDR) reponses to acetylcholine in WT (60.9±6.4% vs 83.9±3.4%, P <0.05) and eET-1 (48.6±5.1% vs 81.5±5.2%, P <0.001). EDR impairment was exaggerated in eET-1 compared to WT ( P <0.05). Meclofenamic acid, an inhibitor of cyclooxygenase, increased EDR in eET-1 compared to WT (78.4±9.4% vs 66.7±3.2%, P <0.01), which was not observed in diabetic mice. L-NAME, an inhibitor of nitric oxide (NO) synthase, completely blocked EDR in WT, eET-1 and diabetic WT, but not in diabetic eET-1 (4.1±1.6%, 6.4±5.7%, 2.2±4.6% and 26.6±4.6%, P <0.05). Apamin plus Tram34, inhibitors of endothelium-dependent hyperpolarization inhibited EDR in the four groups. Endothelium-independent relaxation to sodium nitroprusside, a NO donor, was similar in the four groups. Diabetes reduced media/lumen in WT (2.7±0.3 vs 3.6±0.3, P <0.05) and eET-1 (2.9±0.2 vs 3.8±0.3, P <0.05). Diabetes decreased aortic fibronectin expression in WT (94.0±11.0 vs. 151.9±21.8 RFU/μm 2 , P <0.05) and eET-1 (66.3±8.7 vs. 146.6±20.7 RFU/μm 2 , P <0.05). Conclusion: ET-1 contributes to alterations in several pathways mediating endothelium-dependent relaxation in type-1 diabetes, leading to exaggerated endothelial dysfunction.

scholarly journals Ammonium ion transport by the AMT/Rh homologue LeAMT1;1

2006 ◽  
Vol 396 (3) ◽  
pp. 431-437 ◽  
Author(s):  
Maria Mayer ◽  
Marek Dynowski ◽  
Uwe Ludewig
Keyword(s):  
Structural Data ◽  
Homology Model ◽  
Ammonium Transporter ◽  
Ammonium Ion ◽  
Molecular Architecture ◽  
Gas Channel ◽  
Domains Of Life ◽  
Single Positive ◽  
Key Residues ◽  
Related Proteins

AMT (ammonium transporter)/Rh (Rhesus) ammonium transporters/channels are identified in all domains of life and fulfil contrasting functions related either to ammonium acquisition or excretion. Based on functional and crystallographic high-resolution structural data, it was recently proposed that the bacterial AmtB (ammonium transporter B) is a gas channel for NH3 [Khademi, O'Connell, III, Remis, Robles-Colmenares, Miercke and Stroud (2004) Science 305, 1587–1594; Zheng, Kostrewa, Berneche, Winkler and Li (2004) Proc. Natl. Acad. Sci. U.S.A. 101, 17090–17095]. Key residues, proposed to be crucial for NH3 conduction, and the hydrophobic, but obstructed, pore were conserved in a homology model of LeAMT1;1 from tomato. Transport by LeAMT1;1 was affected by mutations of residues that were predicted to constitute the aromatic recruitment site for NH4+ at the external pore entrance. Despite the structural similarities, LeAMT1;1 was shown to transport only the ion; each transported 14C-methylammonium molecule carried a single positive elementary charge. Similarly, NH4+ (or H+/NH3) was transported, but NH3 conduction was excluded. It is concluded that related proteins and a similar molecular architecture can apparently support contrasting transport mechanisms.

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