Base de dados : MEDLINE
Pesquisa : D01.139.300.050.100 [Categoria DeCS]
Referências encontradas : 4344 [refinar]
Mostrando: 1 .. 10   no formato [Detalhado]

página 1 de 435 ir para página                         

  1 / 4344 MEDLINE  
              next record last record
seleciona
para imprimir
Fotocópia
PubMed Central Texto completo
Texto completo
[PMID]:26864123
[Au] Autor:Soundrarajan N; Cho HS; Ahn B; Choi M; Thong le M; Choi H; Cha SY; Kim JH; Park CK; Seo K; Park C
[Ad] Endereço:Department of Animal Biotechnology, Konkuk University, Seoul 143-701, Korea.
[Ti] Título:Green fluorescent protein as a scaffold for high efficiency production of functional bacteriotoxic proteins in Escherichia coli.
[So] Source:Sci Rep;6:20661, 2016 Feb 11.
[Is] ISSN:2045-2322
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:The availability of simple, robust, and cost-effective methods for the large-scale production of bacteriotoxic peptides such as antimicrobial peptides (AMPs) is essential for basic and pharmaceutical research. However, the production of bacteriotoxic proteins has been difficult due to a high degree of toxicity in bacteria and proteolytic degradation. In this study, we inserted AMPs into the Green fluorescent protein (GFP) in a loop region and expressed them as insoluble proteins in high yield, circumventing the inherent toxicity of AMP production in Escherichia coli. The AMPs inserted were released by cyanogen bromide and purified by chromatography. We showed that highly potent AMPs such as Protegrin-1, PMAP-36, Buforin-2, and Bactridin-1 are produced in high yields and produced AMPs showed similar activities compared to chemically synthesized AMPs. We increased the yield more than two-fold by inserting three copies of Protegrin-1 in the GFP scaffold. The immunogold electron micrographs showed that the expressed Protegrin-1 in the GFP scaffold forms large and small size aggregates in the core region of the inclusion body and become entirely nonfunctional, therefore not influencing the proliferation of E. coli. Our novel method will be applicable for diverse bacteriotoxic peptides which can be exploited in biomedical and pharmaceutical researches.
[Mh] Termos MeSH primário: Peptídeos Catiônicos Antimicrobianos/genética
Escherichia coli/genética
Proteínas de Fluorescência Verde/genética
Corpos de Inclusão/genética
Proteínas/genética
[Mh] Termos MeSH secundário: Peptídeos Catiônicos Antimicrobianos/química
Peptídeos Catiônicos Antimicrobianos/metabolismo
Peptídeos Catiônicos Antimicrobianos/farmacologia
Brometo de Cianogênio/química
Escherichia coli/metabolismo
Escherichia coli/ultraestrutura
Dosagem de Genes
Expressão Gênica
Proteínas de Fluorescência Verde/biossíntese
Corpos de Inclusão/metabolismo
Corpos de Inclusão/ultraestrutura
Testes de Sensibilidade Microbiana
Microscopia Eletrônica de Transmissão
Modelos Moleculares
Agregados Proteicos
Estrutura Secundária de Proteína
Proteínas/química
Proteínas/metabolismo
Proteínas/farmacologia
Pseudomonas aeruginosa/efeitos dos fármacos
Pseudomonas aeruginosa/crescimento & desenvolvimento
Proteínas Recombinantes de Fusão/biossíntese
Proteínas Recombinantes de Fusão/genética
Solubilidade
Staphylococcus aureus/efeitos dos fármacos
Staphylococcus aureus/crescimento & desenvolvimento
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Antimicrobial Cationic Peptides); 0 (PMAP-36); 0 (Protein Aggregates); 0 (Proteins); 0 (Recombinant Fusion Proteins); 0 (buforin II); 0 (protegrin-1); 147336-22-9 (Green Fluorescent Proteins); OS382OHJ8P (Cyanogen Bromide)
[Em] Mês de entrada:1612
[Cu] Atualização por classe:170220
[Lr] Data última revisão:
170220
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160212
[St] Status:MEDLINE
[do] DOI:10.1038/srep20661


  2 / 4344 MEDLINE  
              first record previous record next record last record
seleciona
para imprimir
Fotocópia
Texto completo
[PMID]:26482572
[Au] Autor:Wu XJ; Tang EK; Xu CQ; Yuan ZX
[Ad] Endereço:Integrative Traditional and Western Medicine Hospital of Sichuan Province, Chengdu, Sichuan, China.
[Ti] Título:Hemocompatibility evaluation for peptide fragments of human serum albumin cleaved by cyanogens bromide.
[So] Source:J Biomater Appl;30(7):974-82, 2016 Feb.
[Is] ISSN:1530-8022
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo:We have previously demonstrated that peptide fragments of human serum albumin can be developed into potential renal targeting drug carriers. However, the interactions of these peptide fragments with red blood cells and plasma components are not evaluated well and there is yet no report on the evaluation of the hemocompatibility of peptide fragments. In this study, three kinds of peptide fragments were prepared and identified by amino acid analysis, and the blood compatibility of the peptide fragments was investigated by measuring blood coagulation, platelet and complement activation and hemolysis activity. Results indicated that all the peptide fragments prepared were highly hemocompatible without causing any clot formation, red blood cell aggregation or immune response. In addition, data from the cytotoxicity assay using HeLa cells and Madin-Darby canine kidney cells suggested that these peptide fragments do not induce toxicity towards either cell lines at concentrations up to 5 mg/ml. Therefore, it can be concluded that peptide fragments exhibit good hemocompatibility with no unwanted effect on the viability of renal cells, preliminarily demonstrating that it is safe to use peptide fragments as renal targeting drug carriers.
[Mh] Termos MeSH primário: Brometo de Cianogênio/química
Fragmentos de Peptídeos/química
Albumina Sérica/química
[Mh] Termos MeSH secundário: Animais
Coagulação Sanguínea
Testes de Coagulação Sanguínea
Plaquetas/citologia
Ativação do Complemento
Cães
Portadores de Fármacos
Sistemas de Liberação de Medicamentos
Eritrócitos/efeitos dos fármacos
Células HeLa
Hemólise
Seres Humanos
Rim/citologia
Células Madin Darby de Rim Canino
Plasma/efeitos dos fármacos
Ativação Plaquetária
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Drug Carriers); 0 (Peptide Fragments); 0 (Serum Albumin); OS382OHJ8P (Cyanogen Bromide)
[Em] Mês de entrada:1612
[Cu] Atualização por classe:161230
[Lr] Data última revisão:
161230
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:151021
[St] Status:MEDLINE
[do] DOI:10.1177/0885328215608018


  3 / 4344 MEDLINE  
              first record previous record next record last record
seleciona
para imprimir
Fotocópia
Texto completo
[PMID]:25765310
[Au] Autor:Manoel EA; Dos Santos JC; Freire DM; Rueda N; Fernandez-Lafuente R
[Ad] Endereço:Universidade Federal do Rio de Janeiro, Faculdade de Farmácia, Departamento de Biotecnologia Farmacêutica, Centro de Ciências e Saúde, Bloco B, 1 andar (1 floor), Laboratório Multidisciplinar de Pesquisas em Biotecnologia, CEP 21941902, Rio de Janeiro, Brazil; Department of Biocatalysis, ICP-CSIC, C
[Ti] Título:Immobilization of lipases on hydrophobic supports involves the open form of the enzyme.
[So] Source:Enzyme Microb Technol;71:53-7, 2015 Apr.
[Is] ISSN:1879-0909
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:The lipases from Thermomyces lanuginosus and Pseudomonas cepacia have been immobilized on octyl and cyanogen bromide (CNBr) agarose beads. The immobilization on octyl-agarose is slowed with increasing ionic strength, while the immobilization on CNBr is not significantly affected by the ionic strength. The inhibition of the immobilized preparations with diethyl p-nitrophenylphosphate (D-pNPP) was analyzed. The inhibition was more rapid using octyl-lipase preparations than using covalent preparations, and the covalent preparations were much more sensitive to the reaction medium. The addition of detergent increased the inhibition rate of the covalent preparation while an increase on the ionic strength produced a slowdown of the inhibition rate by D-pNPP for both lipases. The effect of the medium on the activity versus fully soluble substrate (methyl mandelate) was in the same direction. The octyl preparations presented a slight decrease in activity when comparing the results using different concentrations of sodium phosphate buffer (between 0.025 and 1M), while the CNBr preparations suffered drastic drops in its activity at high ionic strength. The results confirm that the lipases immobilized on octyl agarose presented their open form stabilized while the covalent preparation maintains a closing/opening equilibrium that may be modulated by altering the medium.
[Mh] Termos MeSH primário: Enzimas Imobilizadas/metabolismo
Lipase/metabolismo
[Mh] Termos MeSH secundário: Ascomicetos/enzimologia
Proteínas de Bactérias/antagonistas & inibidores
Proteínas de Bactérias/química
Proteínas de Bactérias/metabolismo
Burkholderia cepacia/enzimologia
Brometo de Cianogênio
Enzimas Imobilizadas/antagonistas & inibidores
Enzimas Imobilizadas/química
Proteínas Fúngicas/antagonistas & inibidores
Proteínas Fúngicas/química
Proteínas Fúngicas/metabolismo
Interações Hidrofóbicas e Hidrofílicas
Cinética
Lipase/antagonistas & inibidores
Lipase/química
Concentração Osmolar
Conformação Proteica
Sefarose/análogos & derivados
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Bacterial Proteins); 0 (Enzymes, Immobilized); 0 (Fungal Proteins); 69106-60-1 (octyl-sepharose CL-4B); 9012-36-6 (Sepharose); EC 3.1.1.3 (Lipase); OS382OHJ8P (Cyanogen Bromide)
[Em] Mês de entrada:1512
[Cu] Atualização por classe:150313
[Lr] Data última revisão:
150313
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:150314
[St] Status:MEDLINE


  4 / 4344 MEDLINE  
              first record previous record next record last record
seleciona
para imprimir
Fotocópia
Texto completo
[PMID]:25661879
[Au] Autor:Homaei A
[Ad] Endereço:Department of Biochemistry, Faculty of Sciences, University of Hormozgan, PO Box 3995, Bandar Abbas, Iran. Electronic address: a.homaei@hormozgan.ac.ir.
[Ti] Título:Enhanced activity and stability of papain immobilized on CNBr-activated sepharose.
[So] Source:Int J Biol Macromol;75:373-7, 2015 Apr.
[Is] ISSN:1879-0003
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:Immobilization of papain was carried out by covalent attachment on Sepharose 6B activated by using cyanogen bromide. Immobilization process brought about significant enhancement of storage and thermal stability, stability at extreme pHs, and resistance against the inhibitory effects of various bivalent metal ions with respect to papain. The optimum temperature of papain increased by 20 °C (from 60 to 80 °C) and its optimum pH was shifted from 6.5 to 8.0 upon immobilization. The activation energy of the enzymatic reaction for immobilized papain showed a significant increase as compared with its free form (1.87 kcal mol(-1) K(-1) for free and 4.69 kcal mol(-1) K(-1) for immobilized enzyme). The kinetic parameters, Km and kcat, were estimated to be 0.62 µM and 162 × 10(-4) s(-1) for free and 0.79 µM and 102 × 10(-4) s(-1) for immobilized papain, respectively.
[Mh] Termos MeSH primário: Brometo de Cianogênio/farmacologia
Enzimas Imobilizadas/metabolismo
Papaína/metabolismo
Sefarose/farmacologia
[Mh] Termos MeSH secundário: Biocatálise/efeitos dos fármacos
Estabilidade Enzimática/efeitos dos fármacos
Concentração de Íons de Hidrogênio
Cinética
Reciclagem
Temperatura Ambiente
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Enzymes, Immobilized); 9012-36-6 (Sepharose); EC 3.4.22.2 (Papain); OS382OHJ8P (Cyanogen Bromide)
[Em] Mês de entrada:1512
[Cu] Atualização por classe:150323
[Lr] Data última revisão:
150323
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:150210
[St] Status:MEDLINE


  5 / 4344 MEDLINE  
              first record previous record next record last record
seleciona
para imprimir
Fotocópia
Texto completo
[PMID]:25310875
[Au] Autor:Lee KJ; Lim HS
[Ad] Endereço:Department of Chemistry, Pohang University of Science and Technology (POSTECH) , Pohang 790-784, South Korea.
[Ti] Título:Facile method to sequence cyclic peptides/peptoids via one-pot ring-opening/cleavage reaction.
[So] Source:Org Lett;16(21):5710-3, 2014 Nov 07.
[Is] ISSN:1523-7052
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:A facile method for sequence determination of cyclic peptides/peptoids is described. Macrocyclic peptides/peptoids of 3-10 residues were efficiently synthesized through thioether formation. One-pot reaction of thioether-embedded cyclic peptides/peptoids involving cyanogen bromide-mediated ring-opening and cleavage provides linearized molecules, which can be efficiently sequenced by tandem mass spectrometry.
[Mh] Termos MeSH primário: Brometo de Cianogênio/química
Peptídeos Cíclicos/química
Peptídeos Cíclicos/síntese química
Peptídeos/química
Peptídeos/síntese química
Sulfetos/química
[Mh] Termos MeSH secundário: Sequência de Aminoácidos
Estrutura Molecular
Espectrometria de Massas em Tandem
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T; RESEARCH SUPPORT, U.S. GOV'T, NON-P.H.S.
[Nm] Nome de substância:
0 (Peptides); 0 (Peptides, Cyclic); 0 (Sulfides); OS382OHJ8P (Cyanogen Bromide)
[Em] Mês de entrada:1502
[Cu] Atualização por classe:141107
[Lr] Data última revisão:
141107
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:141014
[St] Status:MEDLINE
[do] DOI:10.1021/ol502788e


  6 / 4344 MEDLINE  
              first record previous record next record last record
seleciona
para imprimir
Fotocópia
PubMed Central Texto completo
Texto completo
[PMID]:25179220
[Au] Autor:Dahal RA; Pramod AB; Sharma B; Krout D; Foster JD; Cha JH; Cao J; Newman AH; Lever JR; Vaughan RA; Henry LK
[Ad] Endereço:From the Department of Basic Sciences, University of North Dakota School of Medicine and Health Sciences, Grand Forks, North Dakota 58203.
[Ti] Título:Computational and biochemical docking of the irreversible cocaine analog RTI 82 directly demonstrates ligand positioning in the dopamine transporter central substrate-binding site.
[So] Source:J Biol Chem;289(43):29712-27, 2014 Oct 24.
[Is] ISSN:1083-351X
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:The dopamine transporter (DAT) functions as a key regulator of dopaminergic neurotransmission via re-uptake of synaptic dopamine (DA). Cocaine binding to DAT blocks this activity and elevates extracellular DA, leading to psychomotor stimulation and addiction, but the mechanisms by which cocaine interacts with DAT and inhibits transport remain incompletely understood. Here, we addressed these questions using computational and biochemical methodologies to localize the binding and adduction sites of the photoactivatable irreversible cocaine analog 3ß-(p-chlorophenyl)tropane-2ß-carboxylic acid, 4'-azido-3'-iodophenylethyl ester ([(125)I]RTI 82). Comparative modeling and small molecule docking indicated that the tropane pharmacophore of RTI 82 was positioned in the central DA active site with an orientation that juxtaposed the aryliodoazide group for cross-linking to rat DAT Phe-319. This prediction was verified by focused methionine substitution of residues flanking this site followed by cyanogen bromide mapping of the [(125)I]RTI 82-labeled mutants and by the substituted cysteine accessibility method protection analyses. These findings provide positive functional evidence linking tropane pharmacophore interaction with the core substrate-binding site and support a competitive mechanism for transport inhibition. This synergistic application of computational and biochemical methodologies overcomes many uncertainties inherent in other approaches and furnishes a schematic framework for elucidating the ligand-protein interactions of other classes of DA transport inhibitors.
[Mh] Termos MeSH primário: Azidas/metabolismo
Cocaína/análogos & derivados
Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo
Simulação de Acoplamento Molecular
[Mh] Termos MeSH secundário: Animais
Azidas/química
Sítios de Ligação
Cocaína/química
Cocaína/metabolismo
Brometo de Cianogênio/metabolismo
Células HeLa
Seres Humanos
Células LLC-PK1
Ligantes
Mesilatos/metabolismo
Simulação de Dinâmica Molecular
Proteínas Mutantes/química
Proteínas Mutantes/metabolismo
Ratos
Especificidade por Substrato
Suínos
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, N.I.H., EXTRAMURAL; RESEARCH SUPPORT, N.I.H., INTRAMURAL
[Nm] Nome de substância:
0 (Azides); 0 (Dopamine Plasma Membrane Transport Proteins); 0 (Ligands); 0 (Mesylates); 0 (Mutant Proteins); 141782-67-4 (RTI 82); 44059-82-7 (methanethiosulfonate); I5Y540LHVR (Cocaine); OS382OHJ8P (Cyanogen Bromide)
[Em] Mês de entrada:1502
[Cu] Atualização por classe:170220
[Lr] Data última revisão:
170220
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:140903
[St] Status:MEDLINE
[do] DOI:10.1074/jbc.M114.571521


  7 / 4344 MEDLINE  
              first record previous record next record last record
seleciona
para imprimir
Fotocópia
Texto completo
[PMID]:24835093
[Au] Autor:dos Santos JC; Garcia-Galan C; Rodrigues RC; de Sant' Ana HB; Gonçalves LR; Fernandez-Lafuente R
[Ad] Endereço:Instituto de Catálisis-CSIC, Campus UAM-CSIC, Cantoblanco, 28049 Madrid, Spain; Departamento de Engenharia Química, Universidade Federal Do Ceará, Campus Do Pici, CEP 60455-760, Fortaleza, CE, Brazil.
[Ti] Título:Improving the catalytic properties of immobilized Lecitase via physical coating with ionic polymers.
[So] Source:Enzyme Microb Technol;60:1-8, 2014 Jun 10.
[Is] ISSN:1879-0909
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Lecitase Ultra has been immobilized on cyanogen bromide agarose (via covalent attachment) and on octyl agarose (via physical adsorption on the hydrophobic support by interfacial activation). Both immobilized preparations have been incubated in dextran sulfate (DS) or polyethylenimine (PEI) solutions to coat the enzyme surface. Then, the activity versus different substrates and under different experimental conditions was evaluated. The PEI coating generally produced a significant increase in enzyme activity, in some cases even by more than a 30-fold factor (using the octyl-Lecitase at pH 5 in the hydrolysis of methyl phenyl acetate). In opposition, the DS coating usually produced some negative effects on the enzyme activity. The rate of irreversible inhibition of the covalent preparation using diethyl p-nitrophenylphosphate did not increase after PEI coating suggesting that the increase in Lecitase activity is not a consequence of the stabilization of the open form of Lecitase. Moreover, the coating greatly increased the stability of the immobilized Lecitase, for example using DS and the covalent preparation, the half-life was increased by a 30-fold factor in 30% acetonitrile. The stabilizing effect was not found in all cases, in certain cases even a certain destabilization is found (e.g., octyl-Lecitase-DS at pH 7). Thus, the effects of the ionic polymer coating strongly depend on the substrate, experimental conditions and immobilization technique employed.
[Mh] Termos MeSH primário: Enzimas Imobilizadas/química
Enzimas Imobilizadas/metabolismo
Fosfolipases A1/química
Fosfolipases A1/metabolismo
[Mh] Termos MeSH secundário: Biotecnologia
Catálise
Materiais Revestidos Biocompatíveis/química
Brometo de Cianogênio
Sulfato de Dextrana
Estabilidade Enzimática
Concentração de Íons de Hidrogênio
Polietilenoimina
Proteínas Recombinantes de Fusão/química
Proteínas Recombinantes de Fusão/metabolismo
Sefarose
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Coated Materials, Biocompatible); 0 (Enzymes, Immobilized); 0 (Recombinant Fusion Proteins); 9002-98-6 (Polyethyleneimine); 9012-36-6 (Sepharose); 9042-14-2 (Dextran Sulfate); EC 3.1.1.32 (Phospholipases A1); OS382OHJ8P (Cyanogen Bromide)
[Em] Mês de entrada:1505
[Cu] Atualização por classe:140519
[Lr] Data última revisão:
140519
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:140520
[St] Status:MEDLINE


  8 / 4344 MEDLINE  
              first record previous record next record last record
seleciona
para imprimir
Fotocópia
Texto completo
[PMID]:24751876
[Au] Autor:Kosicka I; Kristensen T; Bjerring M; Thomsen K; Scavenius C; Enghild JJ; Nielsen NC
[Ad] Endereço:Center for Insoluble Protein Structures (inSPIN), Interdisciplinary Nanoscience Center (iNANO), Aarhus University, Denmark; Department of Chemistry, Aarhus University, Denmark.
[Ti] Título:Preparation of uniformly (13)C,(15)N-labeled recombinant human amylin for solid-state NMR investigation.
[So] Source:Protein Expr Purif;99:119-30, 2014 Jul.
[Is] ISSN:1096-0279
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:A number of diseases are caused by the formation of amyloid fibrils. Detailed understanding of structural features of amyloid fibers is of great importance for our understanding of disease progression and design of agents for diagnostics or potential prevention of protein aggregation. In lack of 3D crystal ordering, solid-state NMR forms the most suited method to determine the structures of the fibrils with atomic resolution. To exploit this potential, large amounts of isotopic-labeled protein need to be obtained through recombinant protein expression. However, expression and purification of amyloidogenic proteins in large amounts remains challenging due to their aggregation potential, toxicity for cells and difficult purification. In this work, we report a method for the production of large amounts of uniformly labeled (13)C,(15)N-human amylin, being one of the most amyloidogenic peptides known. This method utilizes inclusion bodies-directed expression and cheap chemical cleavage with cyanogen bromide in order to minimize the cost of the procedure compared to the use of less efficient proteolytic enzymes. We demonstrate the formation of amylin fibrils in vitro characterized using biophysical methods and electron microscopy, show toxicity towards human cells, and demonstrate that produced material may form the basis for structure determination using solid-state NMR.
[Mh] Termos MeSH primário: Amiloide/química
Polipeptídeo Amiloide das Ilhotas Pancreáticas/isolamento & purificação
Marcação por Isótopo/métodos
[Mh] Termos MeSH secundário: Amiloide/ultraestrutura
Isótopos de Carbono
Dicroísmo Circular
Brometo de Cianogênio/química
Seres Humanos
Polipeptídeo Amiloide das Ilhotas Pancreáticas/química
Isótopos de Nitrogênio
Ressonância Magnética Nuclear Biomolecular/métodos
Proteínas Recombinantes de Fusão/isolamento & purificação
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, NON-U.S. GOV'T
[Nm] Nome de substância:
0 (Amyloid); 0 (Carbon Isotopes); 0 (Islet Amyloid Polypeptide); 0 (Nitrogen Isotopes); 0 (Recombinant Fusion Proteins); OS382OHJ8P (Cyanogen Bromide)
[Em] Mês de entrada:1501
[Cu] Atualização por classe:140602
[Lr] Data última revisão:
140602
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:140423
[St] Status:MEDLINE


  9 / 4344 MEDLINE  
              first record previous record next record last record
seleciona
para imprimir
Fotocópia
PubMed Central Texto completo
Texto completo
[PMID]:24688320
[Au] Autor:Nika H; Hawke DH; Angeletti RH
[Ad] Endereço:Laboratory for Macromolecular Analysis and Proteomics and ; Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, New York 10461, USA; and.
[Ti] Título:N-terminal protein characterization by mass spectrometry after cyanogen bromide cleavage using combined microscale liquid- and solid-phase derivatization.
[So] Source:J Biomol Tech;25(1):19-30, 2014 Apr.
[Is] ISSN:1943-4731
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:A sample preparation method for protein N-terminal peptide isolation from cyanogen bromide (CNBr) protein digests has been developed. In this strategy, the CNBr cleavage was preceded by protein α- and ε-amine acetylation and carboxyamidomethylation in a one-pot reaction scheme. The peptide mixture was adsorbed on ZipTipC18 pipette tips for reaction of the newly generated N-termini with sulfosuccinimidyl-2-(biotinamido) ethyl-1, 3-dithiopropionate. In the subsequent steps, the peptides were exposed in situ to hydroxylamine for reversal of potential hydroxyl group acylation, followed by reductive release of the disulfide-linked biotinamido moiety from the derivatives. The selectively thiol group-functionalized internal and C-terminal peptides were reversibly captured by covalent chromatography on activated thiol-sepharose, leaving the N-terminal fragment in the flow-through fraction. The use of the reversed-phase support as a venue for postcleavage serial modification proved instrumental to ensure throughput and completeness of derivatization. By this sequence of solid-phase reactions, the N-terminal peptide could be recognized uniquely in the MALDI-mass spectra of unfractionated digests by its unaltered mass signature. The use of the sample preparation method was demonstrated with low-picomole amounts of model protein. The N-terminal CNBr fragments were retrieved selectively from the affinity support. The sample preparation method provides for robustness and simplicity of operation using standard equipment available in most biological laboratories and is anticipated to be readily expanded to gel-separated proteins.
[Mh] Termos MeSH primário: Brometo de Cianogênio/química
Fragmentos de Peptídeos/química
Proteínas/química
Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos
[Mh] Termos MeSH secundário: Sequência de Aminoácidos
Cromatografia Líquida de Alta Pressão
Peso Molecular
Peptídeos/síntese química
Peptídeos/química
Proteólise
Sefarose/análogos & derivados
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, N.I.H., EXTRAMURAL
[Nm] Nome de substância:
0 (Peptide Fragments); 0 (Peptides); 0 (Proteins); 69552-82-5 (thiol-sepharose); 9012-36-6 (Sepharose); OS382OHJ8P (Cyanogen Bromide)
[Em] Mês de entrada:1411
[Cu] Atualização por classe:170220
[Lr] Data última revisão:
170220
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:140402
[St] Status:MEDLINE
[do] DOI:10.7171/jbt.14-2501-003


  10 / 4344 MEDLINE  
              first record previous record
seleciona
para imprimir
Fotocópia
PubMed Central Texto completo
Texto completo
[PMID]:24688319
[Au] Autor:Nika H; Hawke DH; Angeletti RH
[Ad] Endereço:Laboratory for Macromolecular Analysis and Proteomics and Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, New York 10461, USA; and.
[Ti] Título:C-terminal protein characterization by mass spectrometry: isolation of C-terminal fragments from cyanogen bromide-cleaved protein.
[So] Source:J Biomol Tech;25(1):1-18, 2014 Apr.
[Is] ISSN:1943-4731
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:A sample preparation method for protein C-terminal peptide isolation from cyanogen bromide (CNBr) digests has been developed. In this strategy, the analyte was reduced and carboxyamidomethylated, followed by CNBr cleavage in a one-pot reaction scheme. The digest was then adsorbed on ZipTipC18 pipette tips for conjugation of the homoserine lactone-terminated peptides with 2,2'-dithiobis (ethylamine) dihydrochloride, followed by reductive release of 2-aminoethanethiol from the derivatives. The thiol-functionalized internal and N-terminal peptides were scavenged on activated thiol sepharose, leaving the C-terminal peptide in the flow-through fraction. The use of reversed-phase supports as a venue for peptide derivatization enabled facile optimization of the individual reaction steps for throughput and completeness of reaction. Reagents were replaced directly on the support, allowing the reactions to proceed at minimal sample loss. By this sequence of solid-phase reactions, the C-terminal peptide could be recognized uniquely in mass spectra of unfractionated digests by its unaltered mass signature. The use of the sample preparation method was demonstrated with low-level amounts of a whole, intact model protein. The C-terminal fragments were retrieved selectively and efficiently from the affinity support. The use of covalent chromatography for C-terminal peptide purification enabled recovery of the depleted material for further chemical and/or enzymatic manipulation. The sample preparation method provides for robustness and simplicity of operation and is anticipated to be expanded to gel-separated proteins and in a scaled-up format to high-throughput protein profiling in complex biological mixtures.
[Mh] Termos MeSH primário: Espectrometria de Massas
Fragmentos de Peptídeos/química
Proteínas/química
[Mh] Termos MeSH secundário: Brometo de Cianogênio/química
Peso Molecular
Proteólise
[Pt] Tipo de publicação:JOURNAL ARTICLE; RESEARCH SUPPORT, N.I.H., EXTRAMURAL
[Nm] Nome de substância:
0 (Peptide Fragments); 0 (Proteins); OS382OHJ8P (Cyanogen Bromide)
[Em] Mês de entrada:1411
[Cu] Atualização por classe:170220
[Lr] Data última revisão:
170220
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:140402
[St] Status:MEDLINE
[do] DOI:10.7171/jbt.14-2501-001



página 1 de 435 ir para página                         
   


Refinar a pesquisa
  Base de dados : MEDLINE Formulário avançado   

    Pesquisar no campo  
1  
2
3
 
           



Search engine: iAH v2.6 powered by WWWISIS

BIREME/OPAS/OMS - Centro Latino-Americano e do Caribe de Informação em Ciências da Saúde