Induction of neutralizing antibodies in mice immunized with an amino-terminal polypeptide of Streptococcus mutans P1 protein produced by a recombinant Bacillus subtilis strain

The oral pathogen Streptococcus mutans expresses a surface protein, P1, which interacts with the salivary pellicle on the tooth surface or with fluid-phase saliva, resulting in bacterial adhesion or aggregation, respectively. P1 is a target of protective immunity. Its N-terminal region has been associated with adhesion and aggregation functions and contains epitopes recognized by efficacious antibodies. In this study, we used Bacillus subtilis, a gram-positive expression host, to produce a recombinant N-terminal polypeptide of P1 (P1(39-512)) derived from the S. mutans strain UA159. Purified P1(39-512) reacted with an anti-full-length P1 antiserum as well as one raised against intact S. mutans cells, indicating preserved antigenicity. Immunization of mice with soluble and heat-denatured P1(39-512) induced antibodies that reacted specifically with native P1 on the surface of S. mutans cells. The anti-P1(39-512) antiserum was as effective at blocking saliva-mediated aggregation of S. mutans cells and better at blocking bacterial adhesion to saliva-coated plastic surfaces compared with the anti-full-length P1 antiserum. In addition, adsorption of the anti-P1 antiserum with P1(39-512) eliminated its ability to block the adhesion of S. mutans cells to abiotic surfaces. The present results indicate that P1(39-512), expressed and purified from a recombinant B. subtilis strain, maintains important immunological features of the native protein and represents an additional tool for the development of anticaries vaccines.

Quantitative control of Lettuce mosaic virus fitness and host defence inhibition by P1-HCPro

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Structure of thrombin complexed with selective non-electrophilic inhibitors having cyclohexyl moieties at P1

The crystal structures of five new non-electrophilic β-strand-templated thrombin active-site inhibitors have been determined bound to the enzyme. Four co-crystallize with hirugen and inhibitor isomorphously to produce thrombin-hirugen crystals (monoclinic, space group C2), while one co-crystallizes in the hexagonal system, space group P65. A 1,4-substituted cyclohexyl moiety is conserved at the P1 position of all the inhibitors, along with a fused hetero-bicyclic five- and six-membered ring that occupies the P2 site. Amino, amidino and aminoimidazole groups are attached to the cyclohexyl ring for recognition at the S1 specificity site, while benzylsulfonyl and diphenyl groups enhance the binding at the S3 subsite. The cyclohexyl groups at the P1 positions of three of the inhibitors appear to be in the energetically favored chair conformation, while the imidazole-substituted cyclohexyl rings are in a boat conformation. Somewhat unexpectedly, the two cyclohexyl-aminoimidazole groups bind differently in the specificity site; the unique binding of one is heretofore unreported. The other inhibitors generally mimic arginyl binding at S1. This group of inhibitors combines the nonelectrophilicity and selectivity of DAPA-like compounds and the more optimal binding features of the S1-S3 sites of thrombin for peptidic molecules, which results in highly potent (binding constants 12 nM-16 pM, one being 1.1 μM) and selective (ranging from 140 to 20 000 times more selective compared with trypsin) inhibitors of thrombin. The binding modes of these novel inhibitors are correlated with their binding constants, as is their selectivity, in order to provide further insight for the design of therapeutic antithrombotic agents that inhibit thrombin directly at the active site.

Influência do fotoperíodo no crescimento e sobrevivência de póslarvas de piracanjuba Brycon orbignyanus (Valenciennes, 1849) (Osteichthyes, Characidae)

This paper aims at studying the influence of photoperiod on the cultivation of Brycon orbignyanus (Valenciennes, 1849) (Osteichthyes, Characidae) post-larvae submitted to four treatments: 0L-24D (L=Light; D=Dark), 10 - L14D, 14L - 10D and 24L-0D, with 3 repetitions. Post-larvae measuring 7.8±0.7mm and weighting 3.5±0.8mg were distributed in 12 aquariums (10L), stocked with 12 post-larvae per aquarium. Fishes were fed daily with Artemia sp. nanplii, in 10 days experiment. A positive relationship between the survival rate (88.9±9.7%) observed in the treatment with 24 hours of luminosity, and the lowest (58.3±8.3%), in the treatment with 24 hours of darkness. No difference was showed (P>0.05) in the mean length and weight of the post-larvae, although there was greater heterogeneity among the post-larvae cultivated at the longer darkness period.

Salmonella enteritidis in the eggs of Japanese quails (coturnix coturnix Japonica - Temminck & Schlegel, 1849) fed diets with different calcium and phosphorus levels

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Ciclo reprodutivo da tabarana, Salminus hilarii (Valenciennes, 1849) (Characidae, Salmininae) na região do baixo rio Sorocaba, SP

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

A Marmota na Corte: recreação e vereda literária no cenário cultural do século XIX (1849-1852)

Pós-graduação em Letras - FCLAS

A poesia lírica na revista Guanabara (1849-1856)

Pós-graduação em Letras - FCLAS

Expressão em Escherichia coli e produção de antissoro policlonal para proteína P1 do Southern bean mosaic virus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Ecomorfologia trófica de Acarichthys heckelii (Müller & Troschel, 1849), Heros efasciatus Heckel, 1840 e Mesonauta insignis (Heckel, 1840) (Perciformes:Cichlidae) nas reservas de desenvolvimento sustentável de Amanã e Mamirauá, Amazônia

Os ciclídeos compõem uma das mais diversas famílias de peixes dulcícolas com 1.900 espécies. Espécies como A. heckelii, H. efasciatus e M. insignis são coletados das reservas de desenvolvimento sustentável de Amanã e Mamirauá de acordo com o projeto de manejo sustentável. Para investigar as variações entre as variáveis morfológicas associadas com a alimentação das espécies para as reservas Amanã e Mamirauá no médio Solimões, foi feita uma análise após eliminar o efeito do tamanho corporal. As três espécies formaram diferentes grupos por espécies, por grupos etários e por ambientes. Os principais caracteres para a formação dos grupos foram a largura da boca e comprimento da cabeça. Diferenças entre os juvenis e os adultos nos atributos área relativa do olho e razão-aspecto da nadadeira caudal foram significativas. A composição da dieta indicou que as três espécies estudadas apresentaram uma convergência alimentar por insetos em ambos os ambientes.

Thiazolide-induced apoptosis in colorectal cancer cells is mediated via the Jun kinase-Bim axis and reveals glutathione-S-transferase P1 as Achilles' heel

Glutathione-S-transferase of the Pi class (GSTP1) is frequently overexpressed in a variety of solid tumors and has been identified as a potential therapeutic target for cancer therapy. GSTP1 is a phase II detoxification enzyme and conjugates the tripeptide glutathione to endogenous metabolites and xenobiotics, thereby limiting the efficacy of antitumor chemotherapeutic treatments. In addition, GSTP1 regulates cellular stress responses and apoptosis by sequestering and inactivating c-Jun N-terminal kinase (JNK). Thiazolides are a novel class of antibiotics for the treatment of intestinal pathogens with no apparent side effects on the host cells and tissue. Here we show that thiazolides induce a GSTP1-dependent and glutathione-enhanced cell death in colorectal tumor cell lines. Downregulation of GSTP1 reduced the apoptotic activity of thiazolides, whereas overexpression enhanced it. Thiazolide treatment caused strong Jun kinase activation and Jun kinase-dependent apoptosis. As a critical downstream target of Jun kinase we identified the pro-apoptotic Bcl-2 homolog Bim. Thiazolides induced Bim expression and activation in a JNK-dependent manner. Downregulation of Bim in turn significantly blocked thiazolide-induced apoptosis. Whereas low concentrations of thiazolides failed to induce apoptosis directly, they potently sensitized colon cancer cells to TNF-related apoptosis-inducing ligand- and chemotherapeutic drug-induced cell death. Although GSTP1 overexpression generally limits chemotherapy and thus antitumor treatment, our study identifies GSTP1 as Achilles' heel and thiazolides as novel interesting apoptosis sensitizer for the treatment of colorectal tumors.