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1.
Sci Rep ; 5: 15958, 2015 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-26514238

RESUMO

We present phonon properties of plutonium metal obtained from a combination of density-functional-theory (DFT) electronic structure and the recently developed compressive sensing lattice dynamics (CSLD). The CSLD model is here trained on DFT total energies of several hundreds of quasi-random atomic configurations for best possible accuracy of the phonon properties. The calculated phonon dispersions compare better with experiment than earlier results obtained from dynamical mean-field theory. The density-functional model of the electronic structure consists of disordered magnetic moments with all relativistic effects and explicit orbital-orbital correlations. The magnetic disorder is approximated in two ways: (i) a special quasi-random structure and (ii) the disordered-local-moment method within the coherent potential approximation. Magnetism in plutonium has been debated intensely, but the present magnetic approach for plutonium is validated by the close agreement between the predicted magnetic form factor and that of recent neutron-scattering experiments.

2.
Neuropharmacology ; 79: 28-36, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24212057

RESUMO

TASK channels belong to the family of K(+) channels with 4 transmembrane segments and 2 pore domains (4TM/2P) per subunit. These channels have been related to apoptosis in cerebellar granule neurons (CGN), as well as cancer in other tissues. TASK current is regulated by hormones, neurotransmitters, anesthetics and divalent cations, which are not selective. Recently, there has been found some organic compounds that inhibit TASK current selectively. In order to find other modulators, we report here a group of five dihydropyrrolo[2,1-a]isoquinolines (DPIs), four of them with putative anticancer activity, that were evaluated on TASK-1 and TASK-3 channels. The compounds 1, 2 and 3 showed IC50 < 320 µM on TASK-1 and TASK-3, intermediate activity on TASK-1/TASK-3 heterodimer, moderate effect over hslo and TREK-1 (500 µM), and practically not inhibition on Shaker-IR, herg and IRK2.1 potassium channels, when they were expressed heterologously in Xenopus laevis oocytes. In rat CGN, 500 µM of these three compounds induced a decrement by >39% of the TASK-carried leak current. Finally, only compound 1 showed significant protection (∼36%) against apoptotic death of CGN induced by K(+) deprivation. These results suggest that DPI compounds could be potential candidates for designing new selective inhibitors of TASK channels.


Assuntos
Isoquinolinas/farmacologia , Proteínas do Tecido Nervoso/antagonistas & inibidores , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio de Domínios Poros em Tandem/antagonistas & inibidores , Pirróis/farmacologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Células Cultivadas , Cerebelo/efeitos dos fármacos , Cerebelo/fisiologia , Canal de Potássio ERG1 , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Canais de Potássio Éter-A-Go-Go/metabolismo , Isoquinolinas/química , Camundongos , Estrutura Molecular , Proteínas do Tecido Nervoso/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Potássio/metabolismo , Bloqueadores dos Canais de Potássio/química , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Canais de Potássio de Domínios Poros em Tandem/genética , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Pirróis/química , Ratos , Ratos Wistar , Superfamília Shaker de Canais de Potássio/metabolismo , Xenopus laevis
3.
Parasitology ; 135(9): 1125-31, 2008 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-18620620

RESUMO

In previous studies, ultrastructural observations revealed a large number of gap junctions (GJs) in the neck and immature proglottid tissues of Taenia solium tapeworms. In these helminths, cytoplasmic glycogen sacs are connected by numerous discrete GJs to other cells throughout the maturing strobilar tissue. Discontinuous sucrose gradients were used to purify membrane fractions containing GJs, which were identified by ultrastructural analysis. A trans-membrane peptide sequence from a highly conserved innexin region was used to construct a 20-amino acid synthetic peptide and used to raise polyclonal antibodies in rabbits that recognized both a 55 and a 67 kDa protein in a Western blot of the GJ-enriched pellet. Immunohistochemistry of larval and adult worm sections incubated with antiserum to the synthetic peptide and a secondary anti-rabbit IgG bound to fluorescein, revealed strong binding to the tegumentary surface of the worm, as well as patchy fluorescent areas in the parenchyma. The results indicate that both the tegument of cysticerci and adult T. solium contain innexin-rich membranes, which may function as a tegumentary transport system for small molecules.


Assuntos
Junções Comunicantes/fisiologia , Proteínas de Insetos/fisiologia , Taenia solium/fisiologia , Sequência de Aminoácidos , Animais , Junções Comunicantes/ultraestrutura , Proteínas de Insetos/genética , Larva , Microscopia Eletrônica , Taenia solium/genética , Taenia solium/ultraestrutura
4.
Curr Top Med Chem ; 8(5): 393-9, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-18393902

RESUMO

This review focuses in the role that antioxidant enzymes play in protection and other important physiological functions such as signal transduction, cell differentiation, growth and apoptosis. Parasites use these enzymes to evade ROS produced by the host immune response and for development inside the host. In the cestoda Taenia solium, three antioxidant enzymes have been studied: a cystosolic Cu,Zn superoxide dismutase that is a target of cestocidal drugs (bencimidazoles); a 2-Cys peroxiredoxin that is a regulatory enzyme of H(2)O(2), molecule essential for several physiological functions; and two isoforms of glutathione transferases that are immunological targets, since they protect immunized mice against cysticercosis. Moreover, all these enzymes are present in all stages of the parasite. These findings suggest that antioxidant enzymes have an important role in T. solium physiology and infection, therefore they might represent the Achilles' heel of the parasite.


Assuntos
Anti-Helmínticos/farmacologia , Antioxidantes/metabolismo , Taenia saginata/efeitos dos fármacos , Taenia saginata/enzimologia , Vacinas/imunologia , Animais , Taenia saginata/imunologia
5.
Rev. bras. pesqui. méd. biol ; Braz. j. med. biol. res;39(3): 365-370, Mar. 2006. graf
Artigo em Inglês | LILACS | ID: lil-421364

RESUMO

We have observed that intracerebroventricular (icv) injection of selective N-methyl-D-aspartic acid (NMDA)-type glutamatergic receptor antagonists inhibits lordosis in ovariectomized (OVX), estrogen-primed rats receiving progesterone or luteinizing hormone-releasing hormone (LHRH). When NMDA was injected into OVX estrogen-primed rats, it induced a significant increase in lordosis. The interaction between LHRH and glutamate was previously explored by us and another groups. The noradrenergic systems have a functional role in the regulation of LHRH release. The purpose of the present study was to explore the interaction between glutamatergic and noradrenergic transmission. The action of prazosin, an alpha1- and alpha2b-noradrenergic antagonist, was studied here by injecting it icv (1.75 and 3.5 µg/6 µL) prior to NMDA administration (1 µg/2 µL) in OVX estrogen-primed Sprague-Dawley rats (240-270 g). Rats manually restrained were injected over a period of 2 min, and tested 1.5 h later. The enhancing effect induced by NMDA on the lordosis/mount ratio at high doses (67.06 ± 3.28, N = 28) when compared to saline controls (6 and 2 µL, 16.59 ± 3.20, N = 27) was abolished by prazosin administration (17.04 ± 5.52, N = 17, and 9.33 ± 3.21, N = 20, P < 0.001 for both doses). Plasma LH levels decreased significantly only with the higher dose of prazosin (1.99 ± 0.24 ng/mL, N = 18, compared to saline-NMDA effect, 5.96 ± 2.01 ng/mL, N = 13, P < 0.05). Behavioral effects seem to be more sensitive to the alpha-blockade than hormonal effects. These findings strongly suggest that the facilitatory effects of NMDA on both lordosis and LH secretion in this model are mediated by alpha-noradrenergic transmission.


Assuntos
Animais , Feminino , Ratos , Antagonistas Adrenérgicos alfa/farmacologia , Hormônio Luteinizante/sangue , Prazosina/farmacologia , Comportamento Sexual Animal/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacos , Injeções Intraventriculares , Hormônio Luteinizante/efeitos dos fármacos , N-Metilaspartato/antagonistas & inibidores , Norepinefrina , Ovariectomia , Postura/fisiologia , Ratos Sprague-Dawley , Comportamento Sexual Animal/fisiologia
6.
Braz J Med Biol Res ; 39(3): 365-70, 2006 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-16501816

RESUMO

We have observed that intracerebroventricular (icv) injection of selective N-methyl-D-aspartic acid (NMDA)-type glutamatergic receptor antagonists inhibits lordosis in ovariectomized (OVX), estrogen-primed rats receiving progesterone or luteinizing hormone-releasing hormone (LHRH). When NMDA was injected into OVX estrogen-primed rats, it induced a significant increase in lordosis. The interaction between LHRH and glutamate was previously explored by us and another groups. The noradrenergic systems have a functional role in the regulation of LHRH release. The purpose of the present study was to explore the interaction between glutamatergic and noradrenergic transmission. The action of prazosin, an alpha1- and alpha2b-noradrenergic antagonist, was studied here by injecting it icv (1.75 and 3.5 microg/6 microL) prior to NMDA administration (1 microg/2 microL) in OVX estrogen-primed Sprague-Dawley rats (240-270 g). Rats manually restrained were injected over a period of 2 min, and tested 1.5 h later. The enhancing effect induced by NMDA on the lordosis/mount ratio at high doses (67.06 +/- 3.28, N = 28) when compared to saline controls (6 and 2 microL, 16.59 +/- 3.20, N = 27) was abolished by prazosin administration (17.04 +/- 5.52, N = 17, and 9.33 +/- 3.21, N = 20, P < 0.001 for both doses). Plasma LH levels decreased significantly only with the higher dose of prazosin (1.99 +/- 0.24 ng/mL, N = 18, compared to saline-NMDA effect, 5.96 +/- 2.01 ng/mL, N = 13, P < 0.05). Behavioral effects seem to be more sensitive to the alpha-blockade than hormonal effects. These findings strongly suggest that the facilitatory effects of NMDA on both lordosis and LH secretion in this model are mediated by alpha-noradrenergic transmission.


Assuntos
Antagonistas Adrenérgicos alfa/farmacologia , Hormônio Luteinizante/sangue , Prazosina/farmacologia , Comportamento Sexual Animal/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacos , Animais , Feminino , Injeções Intraventriculares , Hormônio Luteinizante/efeitos dos fármacos , N-Metilaspartato/antagonistas & inibidores , Norepinefrina/metabolismo , Ovariectomia , Postura/fisiologia , Ratos , Ratos Sprague-Dawley , Comportamento Sexual Animal/fisiologia
7.
Int J Cosmet Sci ; 25(3): 127-35, 2003 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18494894

RESUMO

Axillary malodour is caused by the biotransformation of non-odorous precursors present in apocrine sweat and sebum by the axillary microflora. To counter this, underarm products typically contain high levels of bactericides. However, after an initial decrease in bacterial numbers, the surviving cells grow, producing a concomitant rise in axillary odour. A sustained deodorant effect might be achieved without recourse to bactericidal action if this bacterial growth could be inhibited for extended periods. The current study attempted to inhibit axillary bacterial growth by nutrient deprivation, primarily that of iron (Fe(III)). In vitro analyses identified iron (Fe(III)) as the trace metal whose deprivation had the most profound effect on bacterial growth. Further in vitro investigations with Fe-chelating agents demonstrated that a number of compounds with high binding constants for Fe(III) showed optimal activity. One candidate molecule, diethylenetriaminepentaacetic acid (DTPA), was capable of effectively inhibiting bacterial growth in vitro and on the skin of the lower back. Some bacterial species could additionally utilize iron bound to the iron carrier protein transferrin present in eccrine sweat. This was minimized by the use of an agent, butylated hydroxytoluene (BHT), capable of liberating iron from transferrin via reduction of transferrin-bound ferric ions, allowing subsequent sequestration of Fe(II). Deodorant efficacy evaluation of the combination of DTPA and BHT showed deodorancy benefits over and above that afforded by DTPA alone. This mixture of DTPA and BHT supplemented to a standard ethanolic deodorant, used on 50 people for 2 weeks, was highly effective in limiting bacterial growth in the axilla. Total aerobic bacteria in the axillae were reduced from a mean of log 5.75 (+/-0.73) to log 4.50 (+/-0.90) colony-forming units (cfu) cm(-2) (n = 27, P < 0.01) compared with a non-fortified standard ethanolic deodorant. This was reflected in significant decreases in axillary malodour production, as determined by malodour assessments (P < 0.01). The profile of the axillary microflora was maintained, and all populations were rapidly returned to preuse levels after cessation of product use. This new deodorant technology was benchmarked against leading antimicrobial-based deodorant systems. In three separate deodorant efficacy evaluations, the combination of DTPA and BHT was tested against Triethyl citrate, Triclosan and Farnesol in standard unfragranced ethanolic formulations. The combination of DTPA and BHT showed highly significant deodorancy benefits over and above all these antimicrobial-based deodorant technologies. The combination of an efficient iron chelator with an agent capable of liberating iron from transferrin offers significant benefits in terms of bacterial growth inhibition on the skin and provides a new route to axillary deodorancy.

8.
Proc Natl Acad Sci U S A ; 99(19): 12375-80, 2002 Sep 17.
Artigo em Inglês | MEDLINE | ID: mdl-12218183

RESUMO

The Brucella BvrR/BvrS two-component regulatory system is homologous to the ChvI/ChvG systems of Sinorhizobium meliloti and Agrobacterium tumefaciens necessary for endosymbiosis and pathogenicity in plants. BvrR/BvrS controls cell invasion and intracellular survival. Probing the surface of bvrR and bvrS transposon mutants with monoclonal antibodies showed all described major outer membrane proteins (Omps) but Omp25, a protein known to be involved in Brucella virulence. Absence of Omp25 expression was confirmed by two-dimensional electrophoresis of envelope fractions and by gene reporter studies. The electrophoretic analysis also revealed reduction or absence in the mutants of a second set of protein spots that by matrix-assisted laser desorption ionization MS and peptide mass mapping were identified as a non-previously described Omp (Omp3b). Because bvrR and bvrS mutants are also altered in cell-surface hydrophobicity, permeability, and sensitivity to surface-targeted bactericidal peptides, it is proposed that BvrR/BvrS controls cell envelope changes necessary to transit between extracellular and intracellular environments. A genomic search revealed that Omp25 (Omp3a) and Omp3b belong to a family of Omps of plant and animal cell-associated alpha-Proteobacteria, which includes Rhizobium leguminosarum RopB and A. tumefaciens AopB. Previous work has shown that RopB is not expressed in bacteroids, that AopB is involved in tumorigenesis, and that dysfunction of A. tumefaciens ChvI/ChvG alters surface properties. It is thus proposed that the BvrR/BvrS and Omp3 homologues of the cell-associated alpha-Proteobacteria play a role in bacterial surface control and host cell interactions.


Assuntos
Proteínas da Membrana Bacteriana Externa/genética , Brucella abortus/genética , Brucella abortus/patogenicidade , Genes Bacterianos , Rhizobiaceae/genética , Proteínas da Membrana Bacteriana Externa/isolamento & purificação , Sequência de Bases , DNA Bacteriano/genética , Regulação Bacteriana da Expressão Gênica , Óperon Lac , Dados de Sequência Molecular , Mutação , Filogenia , Especificidade da Espécie , Virulência/genética
9.
J Dent Res ; 81(8): 541-6, 2002 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-12147744

RESUMO

Little is known about the molecular mechanisms that regulate the cementogenesis process, because specific cementum markers are not yet available. To investigate whether a cementoblastoma-conditioned medium-derived protein (CP) could be useful as a cementum biological marker, we studied its expression and distribution in human periodontal tissues, human periodontal ligament, alveolar bone, and cementoblastoma-derived cells. In human periodontal tissues, immunoreactivity to anti-CP was observed throughout the cementoid phase of acellular and cellular cementum, cementoblasts, cementocytes, cells located in the endosteal spaces of human alveolar bone, and in cells in the periodontal ligament located near the blood vessels. Immunopurified CP promoted cell attachment on human periodontal ligament, alveolar bone-derived cells, and gingival fibroblasts. A monoclonal antibody against bovine cementum attachment protein (CAP) cross-reacted with CP. These findings indicate that CP identifies potential cementoblast progenitor cells, is immunologically related to CAP species, and serves as a biological marker for cementum.


Assuntos
Moléculas de Adesão Celular/análise , Cemento Dentário/metabolismo , Tumores Odontogênicos/metabolismo , Adulto , Processo Alveolar/citologia , Processo Alveolar/metabolismo , Análise de Variância , Animais , Anticorpos , Biomarcadores/análise , Bovinos , Adesão Celular , Técnicas de Cultura de Células , Meios de Cultivo Condicionados , Cemento Dentário/citologia , Fibroblastos/citologia , Gengiva/citologia , Gengiva/metabolismo , Humanos , Immunoblotting , Imuno-Histoquímica , Masculino , Tumores Odontogênicos/patologia , Ligamento Periodontal/citologia , Ligamento Periodontal/metabolismo , Estatística como Assunto , Células-Tronco/citologia , Células-Tronco/metabolismo , Células Tumorais Cultivadas
10.
Parasitol Res ; 88(5): 398-404, 2002 May.
Artigo em Inglês | MEDLINE | ID: mdl-12049455

RESUMO

A Taenia solium larval glutathione S-transferase fraction (SGSTF), composed of two proteins with Mr 25,500 (SGSTM1) and 26,500 (SGSTM2), was purified by GSH-sepharose. Its N-terminal sequence analysis revealed that both proteins are related to mammalian mu-class GST enzymes. A cDNA clone coding for SGSTM1 was isolated and the amino acid sequence analysis showed close identity with two Echinococcus GSTs and also high identity with several mu-class GSTs that have been reported. In addition, SGSTM1 presents a similar structure to mu-class GSTs, including the mu loop. The recombinant SGSTM1 is a dimeric protein with enzymatic properties clearly related to mammalian mu-class GSTs. Western blot studies indicated that SGSTM1 is not antigenically related to SGSTM2 or mammalian GSTs from rabbit, pig and rat livers. Immunization with SGSTF and SGSTM2 was highly effective in reducing cysticerci load in murine cysticercosis. In contrast, no protection was obtained using native SGSTM1 and recombinant SGSTM1 as immunogens.


Assuntos
Glutationa Transferase , Taenia solium/enzimologia , Sequência de Aminoácidos , Animais , Clonagem Molecular , Cisticercose/prevenção & controle , DNA de Helmintos/análise , DNA de Helmintos/genética , Feminino , Glutationa Transferase/química , Glutationa Transferase/genética , Glutationa Transferase/imunologia , Glutationa Transferase/metabolismo , Imunização , Isoenzimas , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Análise de Sequência de DNA , Especificidade por Substrato , Taenia solium/genética , Taenia solium/crescimento & desenvolvimento , Taenia solium/imunologia , Vacinas/administração & dosagem
11.
J Org Chem ; 66(12): 4180-6, 2001 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-11397151

RESUMO

A concise access to alpha,beta-dihydroxy alpha-amino acid-derived N-carboxy anhydrides (NCAs) with either like or unlike relative configuration is described. The key steps of the synthetic route are the preparation of the nonracemic 4-alkenyl beta-lactams, through either Horner-type olefination of a common 4-formyl beta-lactam or the Corey-Winter alkene synthesis applied to 4-dihydroxyalkyl beta-lactams, followed by the Sharpless AD reaction, and a subsequent ring expansion of the corresponding 4-substituted 3-hydroxy beta-lactams promoted by TEMPO. The opening of thus-prepared NCAs upon treatment with different O- and N-nucleophiles, including alpha-amino esters which lead to peptides, has also been studied under various reaction conditions.


Assuntos
Aminoácidos/química , Peptídeos/química , beta-Lactamas/química , Conformação Proteica , Análise Espectral
12.
Int J Parasitol ; 30(9): 1007-12, 2000 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-10980291

RESUMO

We isolated and characterised the cDNA that encodes the glycolytic enzyme, triosephosphate isomerase from Taenia solium. A 450 bp DNA fragment was obtained by the polymerase chain reaction using a cDNA from larval stage as template and degenerate oligonucleotides designed from conserved polypeptide sequences from TPIs of several organisms. The fragment was used to screen a T. solium larval stage cDNA library. The isolated cDNA, encoding a protein of 250 amino acids shares 44.8-59.6% positional identity with other known TPIs, in which the catalytic enzyme residues were conserved. The complete coding sequence of the T. solium TPI cDNA was cloned into the expression vector pRSET and expressed as a fusion protein with an N-terminal tail of six histidine residues. The catalytic activity of the purified protein was similar to other TPI enzymes. Northern and Southern blot analysis suggest that in T. solium, single gene exists for triosephosphate isomerase and that the gene is expressed in all stages of the parasite.


Assuntos
Cisticercose/parasitologia , Taenia/genética , Triose-Fosfato Isomerase/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Northern Blotting , Southern Blotting , Cromatografia em Gel , Clonagem Molecular , Primers do DNA/química , DNA de Protozoário/química , Escherichia coli/química , Humanos , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/veterinária , RNA de Protozoário/química , RNA de Protozoário/isolamento & purificação , Proteínas Recombinantes/química , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Suínos , Taenia/enzimologia , Triose-Fosfato Isomerase/química
13.
J Org Chem ; 65(1): 41-6, 2000 Jan 14.
Artigo em Inglês | MEDLINE | ID: mdl-10813893

RESUMO

The synthesis of the (2S,3S,4S)-3,4-dihydroxyhomotyrosine amino acid segment, present in echinocandin B, in its activated form ready for peptide coupling is described. The key steps of the approach are the enantioselective AD reaction of 4-methoxycinnamic acid methyl ester, a completely diastereoselective [2 + 2] hydroxyketene-imine cycloaddition, and the TEMPO-assisted cycloexpansion of the resulting 3-hydroxy beta-lactam to the corresponding alpha-amino acid N-carboxy anhydride (NCA). The smooth opening of the latter upon treatment with L-Thr(OSi(t)BuPh(2))OMe and further acylation with the N-Cbz protected L-4-tert-butyldiphenylsilyloxy proline rendered the southwest portion of echinocandin B.


Assuntos
Aldeídos/química , Antibacterianos/química , Proteínas Fúngicas , Oligopeptídeos/síntese química , Peptídeos Cíclicos , Peptídeos , Aminas/química , Equinocandinas , Análise Espectral
14.
Parasitology ; 117 ( Pt 6): 579-88, 1998 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-9881383

RESUMO

Tapeworms absorb and consume large quantities of glucose through their syncytial tegument, storing the excess as glycogen. Although some studies on the metabolism of glucose in several tapeworms are available, the proteins that mediate its uptake and distribution in their tissue have not been identified. We describe the isolation and characterization of cDNA clones encoding 2 facilitated diffusion glucose transporters (TGTP1 and TGTP2) from Taenia solium, the causal agent of human and porcine cysticercosis. Radio-isotope labelled hexose uptake mediated by TGTP1 expressed in Xenopus oocytes is inhibited by the natural stereoisomers D-glucose and D-mannose but not by L-glucose. Transport by TGTP1 is sensitive to classical inhibitors of facilitated diffusion such as phloretin and cytochalasin B, and insensitive to ouabain. TGTP2 did not function in Xenopus oocytes. Localization studies using specific anti-TGTP1 and anti-TGTP2 antibodies show that TGTP1 is abundant in a number of structures underlying the tegument in adult parasites and larvae, whereas TGTP2 appears to be localized only on the tegumentary surface of the larvae and is not detected in adults.


Assuntos
Proteínas de Helminto/metabolismo , Proteínas de Transporte de Monossacarídeos/metabolismo , Taenia/química , Sequência de Aminoácidos , Animais , Anticorpos Anti-Helmínticos/sangue , Especificidade de Anticorpos , Baculoviridae/genética , Cricetinae , DNA Complementar/genética , Técnica Indireta de Fluorescência para Anticorpo , Vetores Genéticos , Glucose/metabolismo , Proteínas Facilitadoras de Transporte de Glucose , Proteínas de Helminto/química , Manose/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Proteínas de Transporte de Monossacarídeos/química , Proteínas de Transporte de Monossacarídeos/genética , Oócitos/química , Reação em Cadeia da Polimerase , Suínos , Taenia/crescimento & desenvolvimento
15.
Eur J Biochem ; 247(1): 348-55, 1997 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-9249046

RESUMO

We have isolated a cDNA clone of the glycolytic enzyme, triosephosphate isomerase (TPI) from Entamoeba histolytica. Degenerate oligonucleotides obtained by reverse translation of conserved polypeptide sequences, derived from TPIs of other organisms, were used to amplify a 450-bp fragment using E. histolytica cDNA as a template. The fragment was used to screen a cDNA library. The isolated cDNA, encoding a protein of 261 amino acids, shares 43-52.6% positional identity with other known protozoan TPIs. The catalytic residues were conserved; nevertheless, several indels occurred at other regions in the protein sequence. The complete coding sequence of the E. histolytica TPI gene was cloned into the expression vector pRSET and expressed as a wild-type TPI enzyme (E. histolytica TPI) and as a fusion protein with an N-terminal tail of six histidine residues E. histolytica TPI-His6); both recombinant proteins were purified. Molecular modeling of E. histolytica TPI showed an identical topology to the known structures of other TPI molecules, but with a remarkable feature; more than 10 inserted residues are located in the same region of the molecular surface. Studies were performed to detect possible changes that might be caused by the inserted amino acids. The catalytic activity and oligomeric state of the purified protein were similar to that reported for TPI from other sources. In contrast, stability towards dilution, as well as thermal inactivation and unfolding assays, showed that E. histolytica TPI is significantly more stable towards denaturation than Trypanosoma brucei TPI.


Assuntos
Entamoeba histolytica/enzimologia , Triose-Fosfato Isomerase/química , Sequência de Aminoácidos , Animais , Clonagem Molecular , Estabilidade Enzimática , Modelos Moleculares , Dados de Sequência Molecular , Dobramento de Proteína , Triose-Fosfato Isomerase/genética , Triose-Fosfato Isomerase/isolamento & purificação
16.
Eur J Biochem ; 244(3): 700-5, 1997 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-9108237

RESUMO

The gene that encodes for triosephosphate isomerase from Trypanosoma cruzi was cloned and sequenced. In T. cruzi, there is only one gene for triosephosphate isomerase. The enzyme has an identity of 72% and 68% with triosephosphate isomerase from Trypanosoma brucei and Leishmania mexicana, respectively. The active site residues are conserved: out of the 32 residues that conform the interface of dimeric triosephosphate isomerase from T. brucei, 29 are conserved in the T. cruzi enzyme. The enzyme was expressed in Escherichia coli and purified to homogeneity. Data from electrophoretic analysis under denaturing techniques and filtration techniques showed that triosephosphate isomerase from T. cruzi is a homodimer. Some of its structural and kinetic features were determined and compared to those of the purified enzymes from T. brucei and L. mexicana. Its circular dichroism spectrum was almost identical to that of triosephosphate isomerase from T. brucei. Its kinetic properties and pH optima were similar to those of T. brucei and L. mexicana, although the latter exhibited a higher Vmax with glyceraldehyde 3-phosphate as substrate. The sensitivity of the three enzymes to the sulfhydryl reagent methylmethane thiosulfonate (MeSO2-SMe) was determined; the sensitivity of the T. cruzi enzyme was about 40 times and 200 times higher than that of the enzymes from T. brucei and L. mexicana, respectively. Triosephosphate isomerase from T. cruzi and L. mexicana have the three cysteine residues that exist in the T. brucei enzyme (positions 14, 39, 126, using the numbering of the T. brucei enzyme); however, they also have an additional residue (position 117). These data suggest that regardless of the high identity of the three trypanosomatid enzymes, there are structural differences in the disposition of their cysteine residues that account for their different sensitivity to the sulfhydryl reagent. The disposition of the cysteine in triosephosphate isomerase from T. cruzi appears to make it unique for inhibition by modification of its cysteine.


Assuntos
Triose-Fosfato Isomerase/genética , Trypanosoma cruzi/enzimologia , Trypanosoma cruzi/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , DNA Complementar/genética , DNA de Protozoário/genética , Inibidores Enzimáticos/farmacologia , Expressão Gênica , Genes de Protozoários , Cinética , Leishmania mexicana/enzimologia , Leishmania mexicana/genética , Metanossulfonato de Metila/análogos & derivados , Metanossulfonato de Metila/farmacologia , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Homologia de Sequência de Aminoácidos , Especificidade da Espécie , Triose-Fosfato Isomerase/isolamento & purificação , Triose-Fosfato Isomerase/metabolismo , Trypanosoma brucei brucei/enzimologia , Trypanosoma brucei brucei/genética
17.
Int J Parasitol ; 24(2): 265-9, 1994 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-8026905

RESUMO

A glycoprotein-enriched fraction was obtained by Concanavalin A-Sepharose 4B affinity chromatography from a crude extract of T. solium cysticerci. The six most prominent glycoproteins with molecular sizes of 180, 103, 96, 68, 55 and 45 kDa were purified by electro-elution from polyacrylamide gel slices. Ultrastructural localization assays using hyperimmune rabbit sera to each glycoprotein, demonstrated their presence on the tegumentary surface of the bladder wall of T. solium cysticerci. Similar studies showed that the 180 kDa glycoprotein is also present on the surface of the T. solium and T. saginata adult worms, as well as in T. saginata, T. pisiformis and T. crassiceps cysticerci. The 55 kDa glycoprotein, which is one of the most abundant on the cyst surface, was found to correspond to the heavy chain of pig IgG by Western blotting.


Assuntos
Cysticercus/química , Glicoproteínas de Membrana/isolamento & purificação , Animais , Cromatografia de Afinidade , Cysticercus/ultraestrutura , Glicoproteínas de Membrana/análise , Microscopia Imunoeletrônica , Suínos
18.
Arch Med Res ; 25(3): 325-30, 1994.
Artigo em Inglês | MEDLINE | ID: mdl-7803983

RESUMO

The internalization of host macromolecules to the vesicular fluid of T. crassiceps cysticerci was studied in vitro. Uptake of purified class G immunoglobulin was not significantly affected by the specificity of its antigen-recognition site and bovine serum albumin was internalized at a similar rate. Internalization was inhibited at low temperature, being optimal at 37 degrees C and saturation was accomplished only at a protein concentration in the culture medium over 12 mg/ml which is close to the physiological concentration of serum proteins in the host. Morphological studies using markers for adsorptive endocytosis allowed visualization of endocytic vesicles and tracking of their movement across the bladder wall tissue. Degradation of internalized proteins was observed at longer times of incubation, suggesting that proteins are later processed and that degraded host macromolecules can be nutrients for cysticerci. Quantification of this capability of internalization suggests that it might play a role in the in vivo removal of potentially damaging host macromolecules, such as antibodies or complement factors, from the host-parasite interface.


Assuntos
Proteínas Sanguíneas/metabolismo , Cysticercus/metabolismo , Animais , Feminino , Camundongos , Camundongos Endogâmicos BALB C
19.
Mol Biochem Parasitol ; 53(1-2): 17-32, 1992 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-1501637

RESUMO

The complete coding DNA for a Schistosoma mansoni homologue of the epidermal growth factor receptor (SER) was characterized from cDNA clones obtained by homology to the tyrosine kinase domain of erbB. The DNA sequence predicts a 200-kDa translation product that contains a secretory leader, a cysteine-rich extracellular domain, a hydrophobic transmembrane sequence, and an intracellular tyrosine kinase domain. The SER transcript is present in cercariae and adult schistosomes. In addition to SER transcripts, schistosomes produce at least 3 variant transcripts encoding truncated SER products that include the secretory leader and a small portion of the extracellular domain followed by short sequences of unrelated, C-terminal amino acids. Based on these sequences, 2 of the variant mRNAs (class 2 and 5) appear to encode soluble, secreted proteins while one (class 4) encodes an SER variant protein with a hydrophobic C-terminus that may serve as a membrane anchor. Class 2 SER variant transcripts are present at levels comparable to SER transcripts in adult worms but are not detected in cercariae. Class 4 and 5 SER variant transcripts are also found within adult worms but at lower levels. Genomic cloning and characterization demonstrate that the variant SER transcripts arise through alternative splicing of the SER gene.


Assuntos
Receptores ErbB/genética , Splicing de RNA , Schistosoma mansoni/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , DNA/genética , Proteínas de Helminto/genética , Dados de Sequência Molecular , RNA Mensageiro/genética , Homologia de Sequência do Ácido Nucleico , Transcrição Gênica
20.
Rev Esp Cardiol ; 44(4): 283-7, 1991 Apr.
Artigo em Espanhol | MEDLINE | ID: mdl-2068374

RESUMO

We present a 36-year-old woman with a malignant carcinoid syndrome, liver metastases and right heart involvement with severe tricuspid and pulmonic valve stenosis. She was admitted with severe heart failure and poor general status. A combined tricuspid and pulmonic valvuloplasty was performed and she exhibited great clinical improvement that was maintained during the follow-up. We point out the importance of two-dimensional echocardiography in the diagnosis of this pathology and the role of Doppler measurements for the evaluation of the valvuloplasty's results.


Assuntos
Doença Cardíaca Carcinoide/terapia , Cateterismo , Estenose da Valva Pulmonar/terapia , Estenose da Valva Tricúspide/terapia , Adulto , Doença Cardíaca Carcinoide/complicações , Doença Cardíaca Carcinoide/diagnóstico , Feminino , Humanos , Valva Pulmonar , Estenose da Valva Pulmonar/diagnóstico , Estenose da Valva Pulmonar/etiologia , Valva Tricúspide , Estenose da Valva Tricúspide/diagnóstico , Estenose da Valva Tricúspide/etiologia
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