Linearization of a naturally occurring circular protein maintains structure but eliminates hemolytic activity

Cyclotides are a recently discovered family of disulfide rich proteins from plants that contain a circular protein backbone. They are exceptionally stable, as exemplified by their use in native medicine of the prototypic cyclotide kalata B1. The peptide retains uterotonic activity after the plant from which it is derived is boiled to make a medicinal tea. The circular backbone is thought to be in part responsible for the stability of the cyclotides, and to investigate its role in determining structure and biological activity, an acyclic derivative, des-(24-28)-kalata B1, was chemically synthesized and purified. This derivative has five residues removed from the 29-amino acid circular backbone of kalata B1 in a loop region corresponding to a processing site in the biosynthetic precursor protein. Two-dimensional NMR spectra of the peptide were recorded, assigned, and used to identify a series of distance, angle, and hydrogen bonding restraints. These were in turn used to determine a representative family of solution structures. Of particular interest was a determination of the structural similarities and differences between des-(2428)-kalata B1 and native kalata B1. Although the overall three-dimensional fold remains very similar to that of the native circular protein, removal of residues 24-28 of kalata B1 causes disruption of some structural features that are important to the overall stability. Furthermore, loss of hemolytic activity is associated with backbone truncation and linearization.

Nerve growth factor overexpression and autocrine loop in breast cancer cells

We show here that nerve growth factor (NGF), the canonical neurotrophic factor, is synthesized and released by breast cancer cells. High levels of NGF transcript and protein were detected in breast cancer cells by reverse transcription-PCR, Western blotting, ELISA assay and immunohistochemistry. Conversely, NGF production could not be detected in normal breast epithelial cells at either the transcriptional or protein level. Confocal analysis indicated the presence of NGF within classical secretion vesicles. Breast cancer cell-produced NGF was biologically active, as demonstrated by its ability to induce the neuronal differentiation of embryonic neural precursor cells. Importantly, the constitutive growth of breast cancer cells was strongly inhibited by either NGF-neutralizing antibodies or K-252a, a pharmacological inhibitor of NGF receptor TrkA, indicating the existence of an NGF autocrine loop. Together, our data demonstrate the physiological relevance of NGF in breast cancer and its potential interest as a marker and therapeutic target.

Black and white and re(a)d all over again: Indigenous minstrelsy in contemporary Canadian and Australian theatre

This essay considers processes by which community identities are challenged by discussing the use of whiteface as an activist strategy in recent indigenous theatre in Canada and Australia. To understand whiteface, I employ Susan Gubar's notion of racechange, processes that test and even transgress racial borders. I also situate whiteface in relation to the history of blackface minstrelsy. Noting the ways these racial performances affirm the hierarchies of color and how power becomes invested in such color codings, the essay highlights indigenous employment of whiteface as a potential form of critical historiography. I then analyze how whiteface functions in two productions, Daniel David Moses's Almighty Voice and His Wife (1991) in Canada and the Queensland Theatre Company's 2000 revival of George Landen Dann's Fountains Beyond in Australia. My analysis posits that such indigenous performances of whiteface can affirm the identity of the marginalized other even as they destabilize the fixity of race and its meanings.

Epitope map for a growth hormone receptor agonist monoclonal antibody, MAb 263

Monoclonal antibody (MAb) 263 is a widely used monoclonal antibody that recognizes the extracellular domain (ECD) of the GH receptor. It has been shown to act as a GH agonist both in vitro and in vivo, and we report here that it must be divalent to exert its effect on the full-length receptor. To understand the mechanism of its agonist action, we have determined the precise epitope for this antibody using a novel random PCR mutagenesis approach together with expression screening in yeast. A library of 5200 clones of rabbit GH receptor ECD mutants were screened both with MAb 263 and with an anticarboxy-tag antibody to verify complete ECD expression. Sequencing for clones that expressed complete ECD but were not MAb 263 positive identified 20 epitope residues distributed in a discontinuous manner throughout the ECD. The major part of the epitope, as revealed after mapping onto the crystal structure model of the ECD molecule, was located on the side and upper portion of domain 1, particularly within the D - E strand disulfide loop 79 - 96. Molecular dynamics docking of an antibody of the same isotype as MAb 263 was used to dock the bivalent antibody to the 1528-Angstrom(2) epitope and to visualize the likely consequences of MAb binding. The minimized model enables the antibody to grasp two receptors in a pincer-like movement from opposite sides, facilitating alignment of the receptor dimerization domains in a manner similar to, but not identical with, GH.

Abnormalities of the PTH-vitamin D axis and bone turnover markers in children, adolescents and adults with cystic fibrosis: comparison with healthy controls

Abnormalities of calcium and vitamin D metabolism in cystic fibrosis (CF) are well documented. We tested the hypothesis that alterations in calcium metabolism are related to vitamin D deficiency, and that bone resorption is increased relative to accretion in patients with CF. Calcitropic hormones, electrolytes, osteocalcin (OC) and bone alkaline phosphatase (BAP), (markers of bone mineralisation), urinary deoxypyridinoline [total (t) Dpd, a marker of bone resorption] and lumbar spine bone mineral density (LS BMD), expressed as a z-score, were measured in 149 (81 M) CF and 141 (61 M) control children aged 5.3-10.99 years, adolescents aged 11-17.99 years and adults aged 18-55.9 years. Data were analysed by multiple regression to adjust for age. In patients, FEV1% predicted and CRP (as disease severity markers), genotype and pancreatic status (PS) were recorded. The distribution of PTH differed between groups (P

E@D: projeto Serpro de educa????o a dist??ncia

O projeto refere-se ?? experi??ncia do Servi??o Federal de Processamento de Dados (Serpro) na implanta????o de uma solu????o completa de educa????o a dist??ncia, em que, considerando a necessidade de capacita????o de grande contingente de pessoas em curto espa??o de tempo e com baixa disponibilidade de recursos, ocorrida em 1999, surgiu a criatividade de um grupo de t??cnicos para criar uma alternativa que viria a se transformar em solu????o de grande sucesso e de utilidade tanto para o Serpro quanto para seus clientes. Ap??s seis anos do in??cio da experi??ncia, j?? foram desenvolvidos mais de 50 cursos com atendimento a aproximadamente 35 mil pessoas, demonstrando a viabilidade dessa modalidade de aprendizagem na educa????o corporativa

Cheques indevidos: corrigindo um erro de v??rios milh??es de d??lares

O caso trata das dificuldades de tomada de decis??o por parte de uma autoridade federal ao serem constatados erros no repasse de subs??dios para agricultores em um programa governamental. O texto relata o conflito entre as posi????es da auditoria e da ??rea t??cnica a respeito das a????es a serem desenvolvidas; a primeira, considerando a quest??o sob o ponto de vista legal e do interesse p??blico, e a segunda, priorizando o interesse dos agricultores e os impactos pol??ticos de uma decis??o contr??ria ?? categoria. O caso tem sido usado em cursos para subsidiar discuss??es sobre ??tica, accountability, conflito de interesses e gest??o de erros n??o previstos

Emiss??o de certid??o negativa de d??bitos de tributos e contribui????es federais via internet

Obter uma Certid??o Negativa de D??bitos de Tributos e Contribui????es Federais da Pessoa Jur??dica, at?? dezembro de 1997, implicava na presen??a do contribuinte em uma Delegacia da Receita Federal. Com a implanta????o do sistema certid??o negativa na Internet isso mudou; o projeto foi desenvolvido, objetivando permitir que o contribuinte obtenha a Certid??o Negativa sem a necessidade de deslocamento, propiciando-lhe maior conforto e economia financeira, conseq??entemente, diminuindo o fluxo de contribuintes ??s Unidades da SRF. Se o contribuinte est?? em dia com suas obriga????es tribut??rias, ?? poss??vel emiti-la via Internet, no site da SRF. A Certid??o Negativa ?? imprescind??vel para que as empresas participem de licita????es, vendam im??veis ou obtenham cr??dito banc??rio. Pode ficar a cargo da entidade que exige a certid??o, verificar a situa????o fiscal do contribuinte na Internet

Sucess??o gerencial para chefia geral de centros de pesquisa (P&D)

A iniciativa surgiu quando constatada grande defasagem entre o n??vel de qualifica????o do grupo t??cnico-cient??fico (pesquisadores) e do grupo gerencial na EMBRAPA. Foi ent??o implementada nova metodologia de sele????o de chefes das Unidades Descentralizadas. A nova modalidade de sele????o conseguiu combinar participa????o democr??tica dos empregados com a avalia????o objetiva do m??rito dos candidatos e com o compromisso com uma proposta de trabalho compat??vel com os objetivos e a miss??o da UD. Al??m disso, oferece ?? sociedade a oportunidade de participar do gerenciamento de suas unidades de pesquisa

Decreto n?? 6.563: aprova o Estatuto e o Quadro Demonstrativo dos Cargos em Comiss??o e das Fun????es Gratificadas da Funda????o Escola Nacional de Administra????o P??blica - ENAP, e d?? outras provid??ncias.

O PRESIDENTE DA REP??BLICA, no uso das atribui????es que lhe confere o art. 84, incisos IV e VI, al??nea ???a???, da Constitui????o, e tendo em vista o disposto no art. 50 da Lei n. 10.683, de 28 de maio de 2003, decreta: ficam aprovados o Estatuto e o Quadro Demonstrativo dos Cargos em Comiss??o e das Fun????es Gratificadas da Funda????o Escola Nacional de Administra????o P??blica - ENAP, na forma dos Anexos I e II a este Decreto.

??cole Nationale d'Administration Publique: d??velopper les comp??tences des fonctionnaires pour augmenter la capacit?? de L'Etat dans la gestion des politiques publiques

Plus d???une vingtaine d???ann??es d???existence d??di??es ?? la mission de d??velopper les comp??tences des fonctionnaires pour augmenter la capacit?? de l?????tat dans la gestion des politiques publiques fait de l???ENAP un mod??le dans le domaine de la formation. Rattach??e au Minist??re du Plan, du Budget et de la Gestion, l?????cole, d??s sa cr??ation en 1986, a re??ue 245 mille fonctionnaires de tout le pays. En 2007, 26 mille fonctionnaires de l???administration f??d??rale ont attendu un de nos 60 stages offerts.

Lei n?? 8.140, de 28 de dezembro de 1990: altera a denomina????o da Funda????o Centro de Forma????o do Servidor P??blico - FUNCEP e d?? outras provid??ncias

A lei n?? 8.140, de 28 de dezembro de 1990 altera a denomina????o da Funda????o Centro de Forma????o do Servidor P??blico - FUNCEP e d?? outras provid??ncias.