Overexpression of thioredoxin h leads to enhanced activity of starch debranching enzyme (pullulanase) in barley grain

Biochemically active wheat thioredoxin h has been overexpressed in the endosperm of transgenic barley grain. Two DNA constructs containing the wheat thioredoxin h gene (wtrxh) were used for transformation; each contained wtrxh fused to an endosperm-specific B1-hordein promoter either with or without a signal peptide sequence for targeting to the protein body. Twenty-two stable, independently transformed regenerable lines were obtained by selecting with the herbicide bialaphos to test for the presence of the bar herbicide resistance gene on a cotransformed plasmid; all were positive for this gene. The presence of wtrxh was confirmed in 20 lines by PCR analysis, and the identity and level of expression of wheat thioredoxin h was assessed by immunoblots. Although levels varied among the different transgenic events, wheat thioredoxin h was consistently highly expressed (up to 30-fold) in the transgenic grain. Transgenic lines transformed with the B1-hordein promoter with a signal peptide sequence produced a higher level of wheat thioredoxin h on average than those without a signal sequence. The overexpression of thioredoxin h in the endosperm of germinated grain effected up to a 4-fold increase in the activity of the starch debranching enzyme, pullulanase (limit dextrinase), the enzyme that specifically cleaves α-1,6 linkages in starch. These results raise the question of how thioredoxin h enhances the activity of pullulanase because it was found that the inhibitor had become inactive before the enzyme showed appreciable activity.

A strategy for the identification of proteins targeted by thioredoxin

Thioredoxins are 12-kDa proteins functional in the regulation of cellular processes throughout the animal, plant, and microbial kingdoms. Growing evidence with seeds suggests that an h-type of thioredoxin, reduced by NADPH via NADP-thioredoxin reductase, reduces disulfide bonds of target proteins and thereby acts as a wakeup call in germination. A better understanding of the role of thioredoxin in seeds as well as other systems could be achieved if more were known about the target proteins. To this end, we have devised a strategy for the comprehensive identification of proteins targeted by thioredoxin. Tissue extracts incubated with reduced thioredoxin are treated with a fluorescent probe (monobromobimane) to label sulfhydryl groups. The newly labeled proteins are isolated by conventional two-dimensional electrophoresis: (i) nonreducing/reducing or (ii) isoelectric focusing/reducing SDS/PAGE. The isolated proteins are identified by amino acid sequencing. Each electrophoresis system offers an advantage: the first method reveals the specificity of thioredoxin in the reduction of intramolecular vs. intermolecular disulfide bonds, whereas the second method improves the separation of the labeled proteins. By application of both methods to peanut seed extracts, we isolated at least 20 thioredoxin targets and identified 5—three allergens (Ara h2, Ara h3, and Ara h6) and two proteins not known to occur in peanut (desiccation-related and seed maturation protein). These findings open the door to the identification of proteins targeted by thioredoxin in a wide range of systems, thereby enhancing our understanding of its function and extending its technological and medical applications.

Universidades corporativas e vantagens competitivas: um estudo multicaso em franquias brasileiras

O conhecimento tem se mostrado como uma importante fonte de vantagem competitiva para as organizações. Por isso, as empresas buscam, cada vez mais, mecanismos que alavanquem os processos de geração, gestão e disseminação do conhecimento organizacional. Uma das ferramentas, que pode potencializar a transformação do conhecimento em vantagens competitivas, é a universidade corporativa. Nesse contexto, as franquias surgem como um dos objetos de estudos que podem ajudar a explicar esta relação, pois desenvolver e compartilhar conhecimento são aspectos vitais para o franchising. Dessa forma, o principal objetivo deste trabalho é identificar elementos que possam influenciar na geração de vantagens competitivas pela implantação e gestão de universidades corporativas no franchising. Para atingir esses objetivos, foi desenvolvida uma revisão teórica focada nos conceitos de vantagens competitivas, universidades corporativas e franchising. O primeiro esforço mostrou que existe uma lacuna teórica no tema, indicando a necessidade de se realizar um estudo de campo para aprofundar o conhecimento sobre o fenômeno estudado. O estudo de caso, em conjunto com entrevistas em profundidade, foram os métodos escolhidos para realizar o objetivo proposto. Foram realizadas dez entrevistas em profundidade com especialistas em educação corporativa e no franchising, sendo levantadas informações secundárias dos processos de educação do Giraffas e das Universidades Corporativas do Banco do Brasil, Ernest Young do Brasil, Cacau Show e McDonald\'s do Brasil. Para as duas últimas empresas foram desenvolvidos estudos de caso, levantando-se informações comparativas dos seus principais concorrentes. Dessa forma, também foram analisadas informações da Chocolates Brasil, Kopenhagen, Bob\'s e Subway. As principais conclusões do trabalho foram: as universidades corporativas podem impactar de forma expressiva no desenvolvimento de vantagens competitivas no franchising; apesar de as franquias promoverem diversas ações educacionais, as universidades corporativas ajudam a organizar de forma estratégica essas ações, gerando, nos casos estudados, mais impactos que as ações isoladas; as ações que mais geram resultados e barreiras à imitação são as relacionadas à disseminação do conhecimento tácito; a relação entre unidades próprias e franqueadas parece ser importante para se estabelecer estratégias educacionais da rede; a avaliação dos impactos dos treinamentos nos resultados operacionais e financeiros é uma grande dificuldade observada; a literatura preconiza uma relação entre unidades próprias e franqueadas nos estágios de maior maturidade do negócio que não foi observada nos casos estudados; também não foram observadas uma preocupação acentuada com o envolvimento de outros stakeholders e com a busca efetiva de modelos de autofinanciamento.

Description of a College Chamber (manuscript copy), ca. 1744

This humorous, rhyming poem appears to have been co-authored by Thomas Handcock of Massachusetts and Richard Waterman of Warwick, Rhode Island. The document is also signed by Catharine Waterman. Neither of the authors attended Harvard College, and the circumstances of this poem's creation are not known. The poem suggests that they composed the poem while visiting - uninvited - the room of "honest Bob." The poem describes the contents of this college chamber, including the following items: an oak table with a broken leg; paper, a pen, and sand for writing; books, including "Scotch songs," philosophy, Euclid, a book of prayer, Tillotson, and French romances; pipes and tobacco; mugs; a broken violin; copperplate and mezzotint prints; a cat; clothes; two globes; a pair of bellows; a broom; a chamber pot; a candle in a bottle; tea; cups and saucers; a letter to Chloe, to whom the room's inhabitant apparently owed money; a powder horn; a fishing net; a rusty gun; a battledore; a shuttlecock; a cannister; a pair of shoes; and a coffee mill. The poem references events related to the War of Austrian Succession (1740-1748); British Vice Admiral Edward Vernon's siege of Portobello (in present-day Panama) in 1739; the "Rushian War" (perhaps the Russo-Swedish War of 1741-1743); and the War of Jenkins' Ear (the cat in the college chamber, like British Captain Robert Jenkins, has lost an ear).