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1.
Article in English | WPRIM | ID: wpr-880705

ABSTRACT

Adenosine diphosphate (ADP)-ribosylation is a unique post-translational modification that regulates many biological processes, such as DNA damage repair. During DNA repair, ADP-ribosylation needs to be reversed by ADP-ribosylhydrolases. A group of ADP-ribosylhydrolases have a catalytic domain, namely the macrodomain, which is conserved in evolution from prokaryotes to humans. Not all macrodomains remove ADP-ribosylation. One set of macrodomains loses enzymatic activity and only binds to ADP-ribose (ADPR). Here, we summarize the biological functions of these macrodomains in DNA damage repair and compare the structure of enzymatically active and inactive macrodomains. Moreover, small molecular inhibitors have been developed that target macrodomains to suppress DNA damage repair and tumor growth. Macrodomain proteins are also expressed in pathogens, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, these domains may not be directly involved in DNA damage repair in the hosts or pathogens. Instead, they play key roles in pathogen replication. Thus, by targeting macrodomains it may be possible to treat pathogen-induced diseases, such as coronavirus disease 2019 (COVID-19).


Subject(s)
ADP-Ribosylation , COVID-19/metabolism , DNA Repair/physiology , Evolution, Molecular , Humans , Models, Biological , Models, Molecular , N-Glycosyl Hydrolases/metabolism , Poly(ADP-ribose) Polymerases/metabolism , Protein Domains , SARS-CoV-2/pathogenicity
2.
Mem. Inst. Oswaldo Cruz ; 115: e190242, 2020. tab, graf
Article in English | LILACS | ID: biblio-1091241

ABSTRACT

BACKGROUND Ubiquitin (Ub) and Ub-like proteins (Ub-L) are critical regulators of complex cellular processes such as the cell cycle, DNA repair, transcription, chromatin remodeling, signal translation, and protein degradation. Giardia intestinalis possesses an experimentally proven Ub-conjugation system; however, a limited number of enzymes involved in this process were identified using basic local alignment search tool (BLAST). This is due to the limitations of BLAST's ability to identify homologous functional regions when similarity between the sequences dips to < 30%. In addition Ub-Ls and their conjugating enzymes have not been fully elucidated in Giardia. OBJETIVE To identify the enzymes involved in the Ub and Ub-Ls conjugation processes using intelligent systems based on the hidden Markov models (HMMs). METHODS We performed an HMM search of functional Pfam domains found in the key enzymes of these pathways in Giardia's proteome. Each open reading frame identified was analysed by sequence homology, domain architecture, and transcription levels. FINDINGS We identified 118 genes, 106 of which corresponded to the ubiquitination process (Ub, E1, E2, E3, and DUB enzymes). The E3 ligase group was the largest group with 82 members; 71 of which harbored a characteristic RING domain. Four Ub-Ls were identified and the conjugation enzymes for NEDD8 and URM1 were described for first time. The 3D model for Ub-Ls displayed the β-grasp fold typical. Furthermore, our sequence analysis for the corresponding activating enzymes detected the essential motifs required for conjugation. MAIN CONCLUSIONS Our findings highlight the complexity of Giardia's Ub-conjugation system, which is drastically different from that previously reported, and provides evidence for the presence of NEDDylation and URMylation enzymes in the genome and transcriptome of G. intestinalis.


Subject(s)
Ubiquitins/genetics , Giardia lamblia/metabolism , Ubiquitin/genetics , Ubiquitination , Ubiquitins/metabolism , Signal Transduction , Models, Molecular , Giardia lamblia/genetics , Ubiquitin/metabolism
3.
Rev. Assoc. Med. Bras. (1992) ; 65(10): 1283-1289, Oct. 2019. tab, graf
Article in English | LILACS | ID: biblio-1041037

ABSTRACT

SUMMARY Body composition assessment at the molecular level is relevant for the athletic population and its association with high performance is well recognized. The four-compartment molecular model (4C) is the reference method for fat mass (FM) and fat-free mass (FFM) estimation. However, its implementation in a real context is not feasible. Coaches and athletes need practical body composition methods for body composition assessment, and the bioelectrical impedance analysis method (BIA) is usually seen as a useful alternative. The aim of this study was to test the validity of BIA (Tanita, TBF-310) to determine the FM and FFM of elite judo athletes. A total of 29 males were evaluated in a period of weight stability using the reference method (4C) and the alternative method (Tanita, TBF-310). Regarding the 4C method, total-body water was assessed by deuterium dilution, bone mineral by DXA, and body volume by air displacement plethysmography. The slops and intercepts differed from 1 (0.39 and 1.11) and 0 (4.24 and -6.41) for FM and FFM, respectively. FM from Tanita TBF-310 overestimated the 4C method by 0.2 kg although no differences were found for FFM. Tanita TBF-310 explained 21% and 72% respectively in the estimation of absolute values of FM and FFM from the 4C method. Limits of agreement were significant, varying from -6.7 kg to 7.0 kg for FM and from -8.9 kg to 7.5 kg for FFM. In conclusion, TBF-310 Tanita is not a valid alternative method for estimating body composition in highly trained judo athletes.


RESUMO A avaliação da composição corporal ao nível molecular é relevante para a população esportiva e sua associação com o alto rendimento é bem reconhecida. O modelo molecular a quatro compartimentos (4C) é o método de referência para as estimativas de massa gorda (MG) e massa livre de gordura (MLG). No entanto, sua implementação no contexto real não é viável. Técnicos e atletas precisam de métodos práticos de composição corporal para a avaliação da composição corporal e o método de análise de impedância bioelétrica (BIA) é geralmente visto como uma alternativa útil. O objetivo deste estudo foi testar a validade da BIA (Tanita, TBF-310) na determinação de MG e MLG em atletas de elite de judô. Um total de 29 atletas masculinos foi avaliado em um período de estabilidade de peso usando o método de referência (4C) e o método alternativo (Tanita, TBF-310). Em relação ao método a 4C, a água corporal total foi avaliada pela diluição de deutério, mineral ósseo por DXA e volume corporal por pletismografia por deslocamento de ar. Os declives e interceções diferiram de 1 (0,39 e 1,11) e 0 (4,24 e -6,41) para MG e MLG, respectivamente. A MG da Tanita TBF-310 superestimou o método 4C em 0,2 kg, embora não tenham sido encontradas diferenças para MLG. A Tanita TBF-310 explicou 21% e 72%, respectivamente, na estimativa dos valores absolutos de MG e MLG do método a 4C. Os limites de concordância foram grandes, variando de -6,7 kg a 7,0 kg para MG e de -8,9 kg a 7,5 kg para MLG. Em conclusão, a TBF-310 Tanita não é um método alternativo válido para estimar a composição corporal em judocas altamente treinados.


Subject(s)
Humans , Male , Adolescent , Adult , Young Adult , Plethysmography/instrumentation , Body Composition , Models, Molecular , Martial Arts/physiology , Athletes , Reproducibility of Results , Electric Impedance
4.
Chinese Journal of Biotechnology ; (12): 1234-1246, 2019.
Article in Chinese | WPRIM | ID: wpr-771805

ABSTRACT

1,3-1,4-β-glucanase (E.C.3.2.1.73) is an important industrial enzyme which cleave β-glucans into oligosaccharides through strictly cutting the β-1,4 glycosidic bonds in 3-O-substituted glucopyranose units. Microbial 1,3-1,4-β-glucanase belongs to retaining glycosyl hydrolases of family 16 with a jellyroll β-sandwich fold structure. The present paper reviews the industrial application and protein engineering of microbial β-glucanases in the last decades and forecasts the research prospects of microbial β-glucanases.


Subject(s)
Amino Acid Sequence , Glycoside Hydrolases , Models, Molecular , Protein Engineering , Substrate Specificity
5.
Article in Chinese | WPRIM | ID: wpr-775259

ABSTRACT

OBJECTIVE@#To construct a three-dimensional structural model for the selectivity filter in the transient receptor pontential melastatin 8 (TRPM8) ion channel.@*METHODS@#In the Rosetta computational structural biology suite, multiple rounds of modeling with the kinematic loop closure algorithm were performed.@*RESULTS@#After nine rounds of computational modeling, we obtained the models of the selectivity filter within the TRPM8 channel with the lowest energy and high convergence. The model showed that the sidechain of D918 points were away from the central ion permeation pathway, while the sidechains of Q914, D920 and T923 pointed towards it. The glycosylation site N934 was located outside the pore region and its side chain directed to the extracellular water environment.@*CONCLUSIONS@#A three-dimensional structural model for the selectivity filter in the TRPM8 ion channel was constructed, which provides reliable structural information for exploring the mechanism of ion selectivity.


Subject(s)
Algorithms , Ion Channel Gating , Models, Molecular , TRPM Cation Channels , Chemistry
6.
Experimental Neurobiology ; : 658-669, 2019.
Article in English | WPRIM | ID: wpr-785791

ABSTRACT

Anoctamin1 (ANO1) also known as TMEM16A is a transmembrane protein that functions as a Ca²⁺ activated chloride channel. Recently, the structure determination of a fungal Nectria haematococca TMEM16 (nhTMEM16) scramblase by X-ray crystallography and a mouse ANO1 by cryo-electron microscopy has provided the insight in molecular architecture underlying phospholipid scrambling and Ca²⁺ binding. Because the Ca²⁺ binding motif is embedded inside channel protein according to defined structure, it is still unclear how intracellular Ca²⁺ moves to its deep binding pocket effectively. Here we show that EF-hand like region containing multiple acidic amino acids at the N-terminus of ANO1 is a putative site regulating the activity of ANO1 by Ca²⁺ and voltage. The EF-hand like region of ANO1 is highly homologous to the canonical EF hand loop in calmodulin that contains acidic residues in key Ca²⁺-coordinating positions in the canonical EF hand. Indeed, deletion and Ala-substituted mutation of this region resulted in a significant reduction in the response to Ca²⁺ and changes in its key biophysical properties evoked by voltage pulses. Furthermore, only ANO1 and ANO2, and not the other TMEM16 isoforms, contain the EF-hand like region and are activated by Ca²⁺. Moreover, the molecular modeling analysis supports that EF-hand like region could play a key role during Ca²⁺ transfer. Therefore, these findings suggest that EF-hand like region in ANO1 coordinates with Ca²⁺ and modulate the activation by Ca²⁺ and voltage.


Subject(s)
Amino Acids, Acidic , Animals , Calcium , Calmodulin , Chloride Channels , Cryoelectron Microscopy , Crystallography, X-Ray , EF Hand Motifs , Mice , Models, Molecular , Mutagenesis , Nectria , Protein Isoforms
7.
Asian Spine Journal ; : 875-889, 2019.
Article in English | WPRIM | ID: wpr-785500

ABSTRACT

STUDY DESIGN: Development of an in vitro model for assessing the anti-inflammatory efficacies of naringin (Nar) and naringenin (NG).PURPOSE: To evaluate the efficacy of natural flavonoids as therapeutic drugs against anti-inflammatory processes in the nucleus pulposus (NP) cells using in-vitro and in-silico methods.OVERVIEW OF LITERATURE: Intervertebral disc (IVD) disease is a common cause of low back pain. Chronic inflammation and degeneration play a significant role in its etiopathology. Thus, a better understanding of anti-inflammatory agents and their role in IVD degeneration and pro-inflammatory cytokines expression is necessary for pain management and regeneration in IVD.METHODS: We performed primary cell culture of NP cells; immunocytochemistry; gene expression studies of cytokines, metalloproteases, extracellular proteins, and apoptotic markers using quantitative polymerase chain reaction and reverse transcription-polymerase chain reaction (RT-PCR); cytotoxicity assay (MTT); and molecular docking studies using AutoDock 4.2 software (Molecular Graphics Laboratory, La Jolla, CA, USA) to confirm the binding mode of proteins and synthesized complexes. We calculated the mean±standard deviation values and performed analysis of variance and t-test using SPSS ver. 17.0 (SPSS, Inc., Chicago, IL, USA).RESULTS: Molecular docking showed that both Nar and NG bind to the selected genes of interest. Semi-quantitative RT-PCR analysis reveals differential gene expression of collagen (COL)9A1, COL9A2, COL9A3, COL11A2, COMT (catechol-O-methyltransferase), and THBS2 (thrombospondin 2); up regulation of ACAN (aggrecan), COL1A1, COL11A1, interleukin (IL)6, IL10, IL18R1, IL18RAP, metalloprotease (MMP)2, MMP3, MMP9, ADAMTS5 (a disintegrin and metalloproteinase with thrombospondin motifs 5), IGF1R (insulin-like growth factor type 1 receptor), SPARC (secreted protein acidic and cysteine rich), PARK2 (parkin), VDR (vitamin D receptor), and BCL2 (B-cell lymphoma 2); down regulation of IL1A, CASP3 (caspase 3), and nine genes with predetermined concentrations of Nar and NG.CONCLUSIONS: The present study evaluated the anti-inflammatory and regenerative efficiencies of Nar and NG in degenerated human NP cells. Altered gene expressions of cytokines, metalloproteases, extracellular proteins, apoptotic genes were dose responsive. The molecular docking (in silico) studies showed effective binding of these native ligands (Nar and NG) with genes identified as potent inhibitors of inflammation. Thus, these natural flavonoids could serve as anti-inflammatory agents in the treatment of low back pain and sciatica.


Subject(s)
Anti-Inflammatory Agents , Caspase 3 , Collagen , Cysteine , Cytokines , Down-Regulation , Flavonoids , Gene Expression , Humans , Immunohistochemistry , In Vitro Techniques , Inflammation , Interleukin-10 , Interleukins , Intervertebral Disc , Intervertebral Disc Degeneration , Ligands , Low Back Pain , Lymphoma , Metalloproteases , Models, Molecular , Pain Management , Polymerase Chain Reaction , Primary Cell Culture , Regeneration , Sciatica , Thrombospondins , Up-Regulation
8.
Article in English | WPRIM | ID: wpr-719421

ABSTRACT

PURPOSE: The p38 mitogen-activated protein kinase (MAPKs) play a crucial role in the production of pro-inflammatory cytokines and over-expression of it increase cytokines which promote cancer. Among four isoforms, p38α has been well studied in head and neck squamous cell carcinoma (HNSCC) and other cancers as a therapeutic target. p38δ has recently emerged as a potential disease-specific drug target. Elevated serum p38α level in HNSCC was reported earlier from our lab. This study aims to estimate the levels of p38 MAPK-isoforms in the serum of HNSCC and design peptide inhibitor targeting the same. MATERIALS AND METHODS: Levels of p38 MAPK isoforms in the serum of HNSCC and healthy controls were quantified by surface plasmon resonance technology. The peptide inhibitor for p38 MAPK was designed by molecular modeling using Grid-based Ligand Docking with Energetics tools and compared with known specific inhibitors. RESULTS: We have observed highly elevated levels of all four isoforms of p38 MAPK in serum of HNSCC patients compared to the control group. Further, serum p38α, p38β, and p38δ levels were down regulated after therapy in follow-up patients, while p38γ showed no response to the therapy. Present study screened designed peptide WFYH as a specific inhibitor against p38δ. The specific inhibitor of p38δ was found to have no effect on p38α due to great structural difference at ATP binding pocket. CONCLUSION: In this study, first time estimated the levels of p38 MAPK isoforms in the serum of HNSCC. It can be concluded that p38 MAPK isoforms can be a diagnostic and prognostic marker for HNSCC and p38δ as a therapeutic target.


Subject(s)
Adenosine Triphosphate , Carcinoma, Squamous Cell , Cytokines , Epithelial Cells , Follow-Up Studies , Head , Humans , Models, Molecular , Neck , p38 Mitogen-Activated Protein Kinases , Protein Isoforms , Protein Kinases , Surface Plasmon Resonance
9.
Int. j. high dilution res ; 18(3/4): 19-34, 2019.
Article in English | LILACS | ID: biblio-1050037

ABSTRACT

Background: Mercuric chloride is known to inhibit the activity of enzymes. It is used in homeopathy at ultra low concentration (ULC) and is known as Mercurius corrosivus (Merc cor). ULCs of Merc cor are reported to promote enzyme activity. Objective: To see whether the mother tincture (θ) of Merc cor and its ULCs interact with an enzyme invertase at its binding sites and influence enzyme's action on its substrate sucrose. Methods: Merc cor θ (0.15 M HgCl2) was diluted with deionized and distilled (DD) water 1:100 and succussed 10 times to prepare Merc cor 1 cH or 1st potency. This potency was further diluted and succussed in 200 and 1000 steps to prepare 200cH and 1000cH potencies, respectively. Merc cor 200 cH and 1000cH were prepared in 90% ethanol. The two potencies and blank 90% EtOH were diluted with DD water 1:1000 to minimize ethanol content to a negligible amount 0.09%. The control was DD water (0.99g/M). The drugs, EtOH and water control were mixed separately with 0.037 mM invertase in DD water. Using an isothermal calorimetry (ITC) instrument the substrate sucrose (65mM) was injected at 2 µl every 2 min into 300 µl invertase solution 20 times at 25 0C. Molecular modeling study was done to predict possible binding sites and nature of binding between the enzyme and HgCl2, and between the enzyme and water. Potencies after dilution are virtually water. Fluorescence spectra of invertase (4µM) mixed with drug/control solutions were also obtained to see the effect of drugs on protein folding. Results: Thermodynamic parameters like binding constant (K), change in enthalpy(ΔH), entropy(ΔS) and Gibbs free energy(ΔG) showed marked variation in treatment effects on the enzyme. Molecular modeling study also shows variation in binding between invertase and HgCl2, and between invertase and water. Fluorescence spectra show variation in quenching related to different treatments. Conclusion: Merc cor mother tincture and its potencies interact at different binding sites of invertase and modify the enzyme's action on sucrose. So, potencies act as modulators of ligand, sucrose. Drug solutions induce conformational changes in the enzyme. (au)


Subject(s)
Sucrose , Binding Sites , Models, Molecular , Low Potencies , beta-Fructofuranosidase , Homeopathy , Mercuric Chloride
10.
Braz. j. microbiol ; 49(2): 351-357, Apr.-June 2018. graf
Article in English | LILACS | ID: biblio-889245

ABSTRACT

Abstract Economic losses with high mortality rate associated with Porcine circovirus type 2 (PCV2) is reported worldwide. PCV2 commercial vaccine was introduced in 2006 in U.S. and in 2008 in Brazil. Although PCV2 vaccines have been widely used, cases of PCV2 systemic disease have been reported in the last years. Eleven nursery or fattening pigs suffering from PCV2 systemic disease were selected from eight PCV2-vaccinated farms with historical records of PCV2 systemic disease in Southern Brazil. PCV2 genomes were amplified and sequenced from lymph node samples of selected pigs. The comparison among the ORF2 amino acid sequences of PCV2 isolates revealed three amino acid substitutions in the positions F57I, N178S and A190T, respectively. Using molecular modeling, a structural model for the capsid protein of PCV2 was built. Afterwards, the mutated residues positions were identified in the model. The structural analysis of the mutated residues showed that the external residue 190 is close to an important predicted region for antibodies recognition. Therefore, changes in the viral protein conformation might lead to an inefficient antibody binding and this could be a relevant mechanism underlying the recent vaccine failures observed in swine farms in Brazil.


Subject(s)
Animals , Circovirus/chemistry , Capsid Proteins/chemistry , Protein Conformation , Swine , Swine Diseases/virology , Brazil , Models, Molecular , Circovirus/isolation & purification , Circovirus/genetics , Circoviridae Infections/veterinary , Circoviridae Infections/virology , Amino Acid Substitution , Capsid Proteins/genetics
11.
Protein & Cell ; (12): 553-567, 2018.
Article in English | WPRIM | ID: wpr-757973

ABSTRACT

ATP-sensitive potassium channels (K) are energy sensors on the plasma membrane. By sensing the intracellular ADP/ATP ratio of β-cells, pancreatic K channels control insulin release and regulate metabolism at the whole body level. They are implicated in many metabolic disorders and diseases and are therefore important drug targets. Here, we present three structures of pancreatic K channels solved by cryo-electron microscopy (cryo-EM), at resolutions ranging from 4.1 to 4.5 Å. These structures depict the binding site of the antidiabetic drug glibenclamide, indicate how Kir6.2 (inward-rectifying potassium channel 6.2) N-terminus participates in the coupling between the peripheral SUR1 (sulfonylurea receptor 1) subunit and the central Kir6.2 channel, reveal the binding mode of activating nucleotides, and suggest the mechanism of how Mg-ADP binding on nucleotide binding domains (NBDs) drives a conformational change of the SUR1 subunit.


Subject(s)
Adenosine Triphosphate , Metabolism , Amino Acid Sequence , Animals , Binding Sites , Cryoelectron Microscopy , Ligands , Mesocricetus , Mice , Models, Molecular , Nucleotides , Metabolism , Pancreas , Metabolism , Potassium Channels, Inwardly Rectifying , Chemistry , Metabolism , Protein Binding , Protein Multimerization , Protein Structure, Quaternary , Protein Subunits , Chemistry , Metabolism , Sf9 Cells , Spodoptera , Sulfonylurea Receptors , Chemistry , Metabolism
12.
Article in English | WPRIM | ID: wpr-718335

ABSTRACT

BACKGROUND: Molecular genetic abnormalities are observed in over 90% of chronic myelomonocytic leukemia (CMML) cases. Recently, several studies have demonstrated the negative prognostic impact of ASXL1 mutations in CMML patients. We evaluated the prognostic impact of ASXL1 mutations and compared five CMML prognostic models in Korean patients with CMML. METHODS: We analyzed data from 36 of 57 patients diagnosed as having CMML from January 2000 to March 2016. ASXL1 mutation analysis was performed by direct sequencing, and the clinical and laboratory features of patients were compared according to ASXL1 mutation status. RESULTS: ASXL1 mutations were detected in 18 patients (50%). There were no significant differences between the clinical and laboratory characteristics of ASXL1-mutated (ASXL1+) CMML and ASXL1-nonmutated (ASXL1−) CMML patients (all P>0.05). During the median follow-up of 14 months (range, 0–111 months), the overall survival (OS) of ASXL1+ CMML patients was significantly inferior to that of ASXL1− CMML patients with a median survival of 11 months and 19 months, respectively (log-rank P=0.049). An evaluation of OS according to the prognostic models demonstrated inferior survival in patients with a higher risk category according to the Mayo molecular model (log-rank P=0.001); the other scoring systems did not demonstrate a significant association with survival. CONCLUSIONS: We demonstrated that ASXL1 mutations, occurring in half of the Korean CMML patients examined, were associated with inferior survival. ASXL1 mutation status needs to be determined for risk stratification in CMML.


Subject(s)
Follow-Up Studies , Humans , Korea , Leukemia, Myelomonocytic, Chronic , Models, Molecular , Molecular Biology
13.
Protein & Cell ; (12): 629-639, 2018.
Article in English | WPRIM | ID: wpr-756929

ABSTRACT

Mechanosensitive (MS) channels are extensively studied membrane protein for maintaining intracellular homeostasis through translocating solutes and ions across the membrane, but its mechanisms of channel gating and ion selectivity are largely unknown. Here, we identified the YnaI channel as the Na/K cation-selective MS channel and solved its structure at 3.8 Å by cryo-EM single-particle method. YnaI exhibits low conductance among the family of MS channels in E. coli, and shares a similar overall heptamer structure fold with previously studied MscS channels. By combining structural based mutagenesis, quantum mechanical and electrophysiological characterizations, we revealed that ion selective filter formed by seven hydrophobic methionine (YnaI) in the transmembrane pore determined ion selectivity, and both ion selectivity and gating of YnaI channel were affected by accompanying anions in solution. Further quantum simulation and functional validation support that the distinct binding energies with various anions to YnaI facilitate Na/K pass through, which was defined as binding-block mechanism. Our structural and functional studies provided a new perspective for understanding the mechanism of how MS channels select ions driven by mechanical force.


Subject(s)
Cryoelectron Microscopy , Escherichia coli Proteins , Chemistry , Metabolism , Ion Channels , Chemistry , Metabolism , Mechanotransduction, Cellular , Models, Molecular , Quantum Theory
14.
Braz. j. med. biol. res ; 51(3): e7050, 2018. tab, graf
Article in English | LILACS | ID: biblio-889043

ABSTRACT

A new microporous lanthanide metal-organic framework, {[Yb(BTB)(H2O) (DEF)2}n (1, DEF=N,N-Diethylformamide), with 1D nano-sized channels has been constructed by bridging helical chain secondary building units with 1,3,5-benzenetrisbenzoic acid (H3BTB) ligand. Structural characterization suggests that this complex crystallizes in the hexagonal space group P6122 and possesses 1D triangular channels with coordinated water molecules pointing to the channel center. In addition, anti-myocarditis properties of compound 1 were evaluated in vivo. The results showed that compound 1 can improve hemodynamic parameters of, and it may be a good therapeutic option for heart failure in the future.


Subject(s)
Animals , Male , Mice , Anti-Inflammatory Agents/chemistry , Crystallography, X-Ray , Lanthanoid Series Elements/chemistry , Metal-Organic Frameworks/chemistry , Myocarditis/therapy , Anti-Inflammatory Agents/therapeutic use , Metal-Organic Frameworks/therapeutic use , Models, Molecular , Powder Diffraction , Thermogravimetry , X-Ray Diffraction
15.
Mem. Inst. Oswaldo Cruz ; 112(9): 617-625, Sept. 2017. tab, graf
Article in English | LILACS | ID: biblio-894879

ABSTRACT

BACKGROUND Leishmanolysins have been described as important parasite virulence factors because of their roles in the infection of promastigotes and resistance to host's defenses. Leishmania (Viannia) braziliensis contains several leishmanolysin genes in its genome, especially in chromosome 10. However, the functional impact of such diversity is not understood, but may be attributed partially to the lack of structural data for proteins from this parasite. OBJECTIVES This works aims to compare leishmanolysin sequences from L. (V.) braziliensis and to understand how the diversity impacts in their structural and dynamic features. METHODS Leishmanolysin sequences were retrieved from GeneDB. Subsequently, 3D models were built using comparative modeling methods and their dynamical behavior was studied using molecular dynamic simulations. FINDINGS We identified three subgroups of leishmanolysins according to sequence variations. These differences directly affect the electrostatic properties of leishmanolysins and the geometry of their active sites. We identified two levels of structural heterogeneity that might be related to the ability of promastigotes to interact with a broad range of substrates. MAIN CONCLUSION Altogether, the structural plasticity of leishmanolysins may constitute an important evolutionary adaptation rarely explored when considering the virulence of L. (V.) braziliensis parasites.


Subject(s)
Humans , Leishmania braziliensis/genetics , Metalloendopeptidases/genetics , Protein Conformation , Genetic Variation , Models, Molecular
16.
Article in Chinese | WPRIM | ID: wpr-345335

ABSTRACT

<p><b>OBJECTIVE</b>To explore the pathogenesis of protein C deficiency in two pedigrees through mutation detection and model analysis.</p><p><b>METHODS</b>Chromogenic substrate method and enzyme linked immunosorbent assay (ELISA) were used to determine the plasma protein C activity (PC: A) and protein C antigen (PC: Ag) in the two probands and their family members. All of the 9 exons and intron-exon boundaries of the PROC gene were amplified by PCR and analyzed with Sanger sequencing after purification. Corresponding mutate sites of the family members were also amplified and sequenced. The PolyPhen-2 software was used to analyze the perniciousness of the mutations and Clustal X was to analyze the conservatism. The protein model and amino acids interaction of the mutations were analyzed by Swiss-PdbViewer software.</p><p><b>RESULTS</b>The PC: A and PC: Ag of proband 1 was 30% and 35%, while PC:A of his father, mother and aunt were all slightly under the reference range. Two heterozygous missense mutations were found in exons 7 and 5 of the PROC gene, namely c.565 C>T (p.Arg147Trp) and c.383 G>A (p.Gly86Asp). His father and aunt were carriers for c.565 C>T, while his mother had carried c.383 G>A. The PC: A of proband 2 and his son were 50% and 64%, respectively. And they were both positive for p.Arg147Trp. Analysis of PolyPhen-2 indicated that p.Arg147Trp was benign, while p.Gly86Asp was damaging. Clustal X analysis indicated that the p.Arg147Trp was non-conservative, while the p.Gly86Asp was highly conservative. Modeling for the mutant proteins revealed that the simple aromatic ring of Trp147 in p.Arg147Trp destroyed the two hydrogen bonds between Arg147-Lys146 and Arg147-Lys151, and steric hindranted with Arg178. The side chain of Asp86 extended and generated steric clash with Gln90 with the occurrence of p.Gly86Asp. The change of hydrogen bonds and steric effects has altered the spatial configuration of amino acids, which led to unstable mutate proteins and interfered with the secretion.</p><p><b>CONCLUSION</b>Both probands had hereditary protein C deficiencies, for which their parents were all carriers. The heterozygous mutations p.Arg147Trp and p.Gly86Asp were the main cause for PC: A activity decrease. Among these, p.Gly86Asp was discovered for the first time.</p>


Subject(s)
Base Sequence , Child , DNA Mutational Analysis , Methods , Family Health , Female , Heterozygote , Humans , Hydrogen Bonding , Male , Middle Aged , Models, Molecular , Mutation , Pedigree , Phenotype , Protein C , Chemistry , Genetics , Metabolism , Protein C Deficiency , Blood , Genetics , Protein Domains
17.
Protein & Cell ; (12): 590-600, 2017.
Article in English | WPRIM | ID: wpr-756983

ABSTRACT

Entero virus 71 (EV71) causes hand, foot, and mouth disease (HFMD) and occasionally leads to severe neurological complications and even death. Scavenger receptor class B member 2 (SCARB2) is a functional receptor for EV71, that mediates viral attachment, internalization, and uncoating. However, the exact binding site of EV71 on SCARB2 is unknown. In this study, we generated a monoclonal antibody (mAb) that binds to human but not mouse SCARB2. It is named JL2, and it can effectively inhibit EV71 infection of target cells. Using a set of chimeras of human and mouse SCARB2, we identified that the region containing residues 77-113 of human SCARB2 contributes significantly to JL2 binding. The structure of the SCARB2-JL2 complex revealed that JL2 binds to the apical region of SCARB2 involving α-helices 2, 5, and 14. Our results provide new insights into the potential binding sites for EV71 on SCARB2 and the molecular mechanism of EV71 entry.


Subject(s)
Amino Acid Sequence , Animals , Antibodies, Monoclonal , Chemistry , Genetics , Metabolism , Binding Sites , Cell Line , Crystallography, X-Ray , Enterovirus A, Human , Genetics , Allergy and Immunology , Fibroblasts , Virology , Gene Expression , HEK293 Cells , Humans , Immunoglobulin Fab Fragments , Chemistry , Genetics , Metabolism , Lysosome-Associated Membrane Glycoproteins , Chemistry , Genetics , Allergy and Immunology , Mice , Models, Molecular , Protein Binding , Protein Conformation, alpha-Helical , Protein Conformation, beta-Strand , Protein Interaction Domains and Motifs , Receptors, Scavenger , Chemistry , Genetics , Allergy and Immunology , Receptors, Virus , Chemistry , Genetics , Allergy and Immunology , Recombinant Fusion Proteins , Chemistry , Genetics , Allergy and Immunology , Sequence Alignment , Sequence Homology, Amino Acid , Sf9 Cells , Spodoptera , Thermodynamics
18.
Cell Journal [Yakhteh]. 2017; 19 (1): 65-83
in English | IMEMR | ID: emr-185794

ABSTRACT

Objective: Druggability of a target protein depends on the interacting micro-environment between the target protein and drugs. Therefore, a precise knowledge of the interacting micro-environment between the target protein and drugs is requisite for drug discovery process. To understand such micro-environment, we performed in silico interaction analysis between a human target protein, Dipeptidyl Peptidase-IV [DPP-4], and three anti-diabetic drugs [saxagliptin, linagliptin and vildagliptin]


Materials and Methods: During the theoretical and bioinformatics analysis of micro-environmental properties, we performed drug-likeness study, protein active site predictions, docking analysis and residual interactions with the protein-drug interface. Micro-environmental landscape properties were evaluated through various parameters such as binding energy, intermolecular energy, electrostatic energy, van der Waals'+H-bond+desolvo energy [EVHD] and ligand efficiency [LE] using different in silico methods. For this study, we have used several servers and software, such as Molsoft prediction server, CASTp server, AutoDock software and LIGPLOT server


Results: Through micro-environmental study, highest log P value was observed for linagliptin [1.07]. Lowest binding energy was also observed for linagliptin with DPP-4 in the binding plot. We also identified the number of H-bonds and residues involved in the hydrophobic interactions between the DPP-4 and the anti-diabetic drugs. During interaction, two H-bonds and nine residues, two H-bonds and eleven residues as well as four H-bonds and nine residues were found between the saxagliptin, linagliptin as well as vildagliptin cases and DPP-4, respectively


Conclusion: Our in silico data obtained for drug-target interactions and micro-environmental signature demonstrates linagliptin as the most stable interacting drug among the tested anti-diabetic medicines


Subject(s)
Humans , Molecular Targeted Therapy , Protein Binding , Drug Discovery , Dipeptidyl Peptidase 4/drug effects , Dipeptidyl-Peptidase IV Inhibitors , Models, Molecular
19.
Braz. j. microbiol ; 47(1): 251-258, Jan.-Mar. 2016. tab, graf
Article in English | LILACS | ID: lil-775122

ABSTRACT

Abstract Dengue is a major worldwide public health problem, especially in the tropical and subtropical regions of the world. Primary infection with a single Dengue virus serotype causes a mild, self-limiting febrile illness called dengue fever. However, a subset of patients who experience secondary infection with a different serotype can progress to a more severe form of the disease, called dengue hemorrhagic fever. The four Dengue virus serotypes (1–4) are antigenically and genetically distinct and each serotype is composed of multiple genotypes. In this study we isolated one Dengue virus 1 serotype, named BR/Alfenas/2012, from a patient with dengue hemorrhagic fever in Alfenas, South Minas Gerais, Brazil and molecular identification was performed based on the analysis of NS5 gene. Swiss mice were infected with this isolate to verify its potential to induce histopathological alterations characteristic of dengue. Liver histopathological analysis of infected animals showed the presence of inflammatory infiltrates, hepatic steatosis, as well as edema, hemorrhage and necrosis focal points. Phylogenetic and evolutionary analyses based on the envelope gene provided evidence that the isolate BR/Alfenas/2012 belongs to genotype V, lineage I and it is probably derived from isolates of Rio de Janeiro, Brazil. The isolate BR/Alfenas/2012 showed two unique amino acids substitutions (SER222THRE and PHE306SER) when compared to other Brazilian isolates from the same genotype/lineage. Molecular models were generated for the envelope protein indicating that the amino acid alteration PHE 306 SER could contribute to a different folding in this region located within the domain III. Further genetic and animal model studies using BR/Alfenas/2012 and other isolates belonging to the same lineage/genotype could help determine the relation of these genetic alterations and dengue hemorrhagic fever in a susceptible population.


Subject(s)
Animals , Humans , Mice , Dengue Virus/classification , Dengue Virus/genetics , Dengue/virology , Genetic Variation , Genotype , Phylogeny , Amino Acid Substitution , Animal Structures/pathology , Brazil , Disease Models, Animal , Dengue Virus/isolation & purification , Gene Products, env/chemistry , Gene Products, env/genetics , Histocytochemistry , Microscopy , Models, Molecular , Point Mutation , Protein Conformation , Viral Nonstructural Proteins/genetics
20.
Protein & Cell ; (12): 792-803, 2016.
Article in English | WPRIM | ID: wpr-757369

ABSTRACT

MRG proteins are conserved during evolution in fungi, flies, mammals and plants, and they can exhibit diversified functions. The animal MRGs were found to form various complexes to activate gene expression. Plant MRG1/2 and MRG702 were reported to be involved in the regulation of flowering time via binding to H3K36me3-marked flowering genes. Herein, we determined the crystal structure of MRG701 chromodomain (MRG701). MRG701 forms a novel dimerization fold both in crystal and in solution. Moreover, we found that the dimerization of MRG chromodomains is conserved in green plants. Our findings may provide new insights into the mechanism of MRGs in regulation of gene expression in green plants.


Subject(s)
Amino Acid Sequence , Arabidopsis , Genetics , Metabolism , Arabidopsis Proteins , Chemistry , Genetics , Metabolism , Binding Sites , Chromosomal Proteins, Non-Histone , Chemistry , Genetics , Metabolism , Cloning, Molecular , Crystallography, X-Ray , Escherichia coli , Genetics , Metabolism , Gene Expression , Histones , Chemistry , Genetics , Metabolism , Models, Molecular , Oryza , Genetics , Metabolism , Peptides , Chemistry , Genetics , Metabolism , Protein Binding , Protein Interaction Domains and Motifs , Protein Isoforms , Chemistry , Genetics , Metabolism , Protein Multimerization , Protein Structure, Secondary , Recombinant Proteins , Chemistry , Genetics , Metabolism , Sequence Alignment , Sequence Homology, Amino Acid , Viridiplantae , Genetics , Metabolism
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