Mechanisms of acetohydroxyacid synthases

Acetohydroxyacid synthases are thiamin diphosphate- (ThDP-) dependent biosynthetic enzymes found in all autotrophic organisms. Over the past 4-5 years, their mechanisms have been clarified and illuminated by protein crystallography, engineered mutagenesis and detailed single-step kinetic analysis. Pairs of catalytic subunits form an intimate dimer containing two active sites, each of which lies across a dimer interface and involves both monomers. The ThDP adducts of pyruvate, acetaldehyde and the product acetohydroxyacids can be detected quantitatively after rapid quenching. Determination of the distribution of intermediates by NMR then makes it possible to calculate individual forward unimolecular rate constants. The enzyme is the target of several herbicides and structures of inhibitor-enzyme complexes explain the herbicide-enzyme interaction.

Acetohydroxyacid synthase and its role in the biosynthetic pathway for branched-chain amino acids

The branched-chain amino acids are synthesized by plants, fungi and microorganisms, but not by animals. Therefore, the enzymes of this pathway are potential target sites for the development of antifungal agents, antimicrobials and herbicides. Most research has focused upon the first enzyme in this biosynthetic pathway, acetohydroxyacid synthase (AHAS) largely because it is the target site for many commercial herbicides. In this review we provide a brief overview of the important properties of each enzyme within the pathway and a detailed summary of the most recent AHAS research, against the perspective of work that has been carried out over the past 50 years.

Defenses against oxidative stress in Neisseria gonorrhoeae: A system tailored for a challenging environment

Neisseria gonorrhoeae is a host-adapted pathogen that colonizes primarily the human genitourinary tract. This bacterium encounters reactive oxygen and reactive nitrogen species as a consequence of localized inflammatory responses in the urethra of males and endocervix of females and also of the activity of commensal lactobacilli in the vaginal flora. This review describes recent advances in the understanding of defense systems against oxidative stress in N. gonorrhoeae and shows that while some of its defenses have similarities to the paradigm established with Escherichia coli, there are also some key differences. These differences include the presence of a defense system against superoxide based on manganese ions and a glutathione-dependent system for defense against nitric oxide which is under the control of a novel MerR-like transcriptional regulator. An understanding of the defenses against oxidative stress in N. gonorrhoeae and their regulation may provide new insights into the ways in which this bacterium survives challenges from polymorphonuclear leukocytes and urogenital epithelial cells.

Sequence analysis of a CTX-M-1 IncI1 plasmid found in Salmonella enterica 4,5,12,i:-, Escherichia coli and Klebsiella pneumoniae on a single UK pig farm

OBJECTIVES: In 2009, CTX-M Enterobacteriaceae and Salmonella isolates were recovered from a UK pig farm, prompting studies into the dissemination of the resistance and to establish any relationships between the isolates. METHODS: PFGE was used to elucidate clonal relationships between isolates whilst plasmid profiling, restriction analysis, sequencing and PCR were used to characterize the CTX-M-harbouring plasmids. RESULTS: Escherichia coli, Klebsiella pneumoniae and Salmonella 4,5,12:i:- and Bovismorbificans resistant to cefotaxime (n = 65) were recovered and 63 were shown by PCR to harbour a group 1 CTX-M gene. The harbouring hosts were diverse, but the group 1 CTX-M plasmids were common. Three sequenced CTX-M plasmids from E. coli, K. pneumoniae and Salmonella enterica serotype 4,5,12:i:- were identical except for seven mutations and highly similar to IncI1 plasmid ColIb-P9. Two antimicrobial resistance regions were identified: one inserted upstream of yacABC harbouring ISCR2 transposases, sul2 and floR; and the other inserted within shfB of the pilV shufflon harbouring the ISEcp1 transposase followed by blaCTX-M-1. CONCLUSIONS: These data suggest that an ST108 IncI1 plasmid encoding a blaCTX-M-1 gene had disseminated across multiple genera on this farm, an example of horizontal gene transfer of the blaCTX-M-1 gene.

Receptor binding studies disclose a novel class of high-affinity inhibitors of the Escherichia coli FimH adhesin

Mannose-binding type 1 pili are important virulence factors for the establishment of Escherichia coli urinary tract infections (UTIs). These infections are initiated by adhesion of uropathogenic E. coli to uroplakin receptors in the uroepithelium via the FimH adhesin located at the tips of type 1 pili. Blocking of bacterial adhesion is able to prevent infection. Here, we provide for the first time binding data of the molecular events underlying type 1 fimbrial adherence, by crystallographic analyses of the FimH receptor binding domains from a uropathogenic and a K-12 strain, and affinity measurements with mannose, common mono- and disaccharides, and a series of alkyl and aryl mannosides. Our results illustrate that the lectin domain of the FimH adhesin is a stable and functional entity and that an exogenous butyl alpha- D-mannoside, bound in the crystal structures, exhibits a significantly better affinity for FimH (K-d = 0.15 muM) than mannose (K-d = 2.3 muM). Exploration of the binding affinities of alpha-D-mannosides with longer alkyl tails revealed affinities up to 5 nM. Aryl mannosides and fructose can also bind with high affinities to the FimH lectin domain, with a 100-fold improvement and 15-fold reduction in affinity, respectively, compared with mannose. Taken together, these relative FimH affinities correlate exceptionally well with the relative concentrations of the same glycans needed for the inhibition of adherence of type 1 piliated E. coli. We foresee that our findings will spark new ideas and initiatives for the development of UTI vaccines and anti-adhesive drugs to prevent anticipated and recurrent UTIs.

Engineering modular viral scaffolds for targeted bacterial population editing

Bacteria are central to human health and disease, but existing tools to edit microbial consortia are limited. For example, broad-spectrum antibiotics are unable to precisely manipulate bacterial communities. Bacteriophages can provide highly specific targeting of bacteria, but assembling well-defined phage cocktails solely with natural phages can be a time-, labor- and cost-intensive process. Here, we present a synthetic biology strategy to modulate phage host ranges by engineering phage genomes in Saccharomyces cerevisiae. We used this technology to redirect Escherichia coli phage scaffolds to target pathogenic Yersinia and Klebsiella bacteria, and conversely, Klebsiella phage scaffolds to target E. coli by modular swapping of phage tail components. The synthetic phages achieved efficient killing of their new target bacteria and were used to selectively remove bacteria from multi-species bacterial communities with cocktails based on common viral scaffolds. We envision this approach accelerating phage biology studies and enabling new technologies for bacterial population editing.

Early intervention for conversion disorder: neurologists and psychiatrists working together

It has been shown that for the reaction catalyzed by "biodegradative" L-threonine dehydratase from E. coli strains K-12 and 980 in 0.5 M phosphate-carbonate buffer, pH 8.4 and pH 9.5, the plots of initial reaction rate (v) versus the initial substrate concentration ([S]0 are characterized by several inflection points, i. e. an intermediate plateau. The plot of v versus the allosteric activator (AMP) concentration have very complicated shapes: there are several inflection points, and also the maximum at L-threonine concentration equal to 3-10(2) and 5-10(-2) M. High AMP concentrations inhibit the enzyme at high substrate concentrations. The reduced glutathion dose not influence the enzyme and does not alter the activating effect of AMP. On the basis of the data obtained it is proposed that the substrate and AMP shift the equilibrium between multiple oligomeric enzyme forms differing in catalytic activity and kinetic manifestations of allosteric interactions between the active and allosteric AMP-binding sites towards polymerization. Thus, the functioning the enzyme under study is discussed in the frames of the model of dissociating regulatory enzymes with multiple intermediate oligomeric forms.

Expresión genética en escherichia coli controlada por medio de temperatura y potencialmente con luz para el dise��o de agentes terapéuticos

Utilizando la interfase entre la biología sintética y la nanotecnología, se propone el dise��o de un material (bacteria), con sensibilidad optotérmica; es decir, que la sinergia entre la capacidad de las nanopartículas metálicas para convertir la luz absorbida en calor y la propiedad de respuesta térmica del termómetro de RNA, permitan una respuesta a la exposición de longitudes de onda concretas a nivel local, y además puedan ser controlados con el uso de incrementos globales en la temperatura del sistema, permitiendo con ello un precursor viable para el dise��o posterior de nuevos agentes terapéuticos que actúen de forma específica y controlada. La metodología para construir los plásmidos incluyó métodos clásicos para su clonación (digestión, ligación y transformación), para la biosíntesis de AgNPs se trabajó con diferentes concentraciones de AgNO3, y cuatro cepas de Escherichia coli en fase log. Se comprobó la viabilidad de las cepas por medio de la técnica de la gota en superficie, así como la actividad del termosensor y expresión de la prote��na mcherry.Contribuciones y Conclusiones: Se realizaron satisfactoriamente las transformaciones de Escherichia coli k

Avaliação dos teores de N, P, K, Ca, Mg e S em plantas de gravioleira cultivadas em solução nutritiva com omissão de macronutrientes.

Com o objetivo de avaliar os teores de N, P, K, Ca, Mg e S nas folhas, caule e raízes de plantar de gravioleira, conduziu-se experimento em casa de vegetação, utilizando a técnica do elemento faltante. O delineamento experimental foi inteiramente casualizado com quatro repetições e os tratamentos foram: completo, omissão de N, P, K, Ca, Mg e S. Os resultados analíticos evidenciaram que a omissão individual de macronutrientes promoveu alterações nas concentrações de N, P, K, Ca, Mg e S nas plantar de gravioleira. Foram verificados efeitos de varias interações entre os nutrientes. Com base nos teores em g kg-1 dos macronutrientes nas folhas, infere-se, em uma primeira aproximação, dos valores adequados (completo) e do deficiente (omissão), sendo: N =14,70 e 8,82; P = 0,92 e 0,47; K = 12,35 e 2,62; Ca = 14,11 e 3,44; Mg = 3,59 e 1,09; S = 5,32 e 2,30.

Virtual globes as a platform for developing spatial literacy

Dissertation submitted in partial fulfilment of the requirements for the Degree of Master of Science in Geospatial Technologies

Nutrição mineral de hortaliças LXX: acumulação de matéria seca e recrutamento de macronutrientes pelo melão (Cucimis Melo L. cv. Valenciano Amarelo CAC) cultivado em latossolo vermelho amarelo em presidente Venceslau, SP

Com o objetivo de determinar: - a curva de crescimento do melão (Cucumis melo L.), através da acumulação de matéria seca; - os teores e consequentes acúmulos de nutrientes nos órgãos aéreos da planta, em diferentes estádios de crescimento; - a exportação de nutrientes na colheita de frutos, no ponto de consumo. Foi conduzido um ensaio em condições de campo o qual consistiu da amostragem em cinco estádios de crescimento - 15, 30, 45, 60 e 75 dias após a emergência - de plantas competitivas, as quais eram cortadas rente ao solo, divididas em caule e ramos, folhas, flores e frutos, para determinação de quantidade de matéria seca e análise química para macronutrientes minerais. Os tratamentos, então representados por épocas de amostragem, constaram de um delineamento em blocos casualizados com quatro repetições. Concluiu-se que: - o crescimento das plantas é inicialmente lento, intensificando-se posteriormente, com maiores incrementos entre 30 e 45 dias; - no final do ciclo, a planta acumulou 905,88 g de matéria seca, assim distribuída; caule e ramos 19,38%, folhas 30,32% e flores e frutos 50,30%; - a concentração dos nutrientes na parte aérea variou com a idade e o órgão considerado; - verifica-se nos frutos uma tendência de redução nos teores dos macronutrientes (com exce��ão do enxofre); - a parte aérea acumulou, aos 75 dias, em miligramas por planta: 23,08 de N; 3,46 de P; 28,90 de K; 12,74 de Ca; 5,55 de Mg; 1,59 de S. - são exportados numa colheita de uma tonelada de frutos, cerca de 1,78 kg de N; 0,33 kg de P; 2,65 kg de K; 0,14 kg de Ca; 0,21 kg de Mg; 0,09 kg de S.

Acúmulo de macronutrientes pela bananeira cv. prata em diferentes estádios de desenvolvimento

Visando estabelecer a marcha de absorção, quantidade exportada e reciclada de nitrogênio, fósforo, potássio e cálcio na bananeira cultivar Prata, instalou-se um ensaio no Estado do Espírito Santo, em janeiro de 1980, em terreno com declividade média de 40% e solo cambissólico distrófico. Sorteou-se três plantas matrizes do plantio até 465 dias após (colheita),sendo bimensal até 300 dias e mensal até o final, totalizando doze estádios de desenvolvimento, e três plantas rebento, mensalmente, da emissão até 300 dias após, atingindo nove estádios. Cada bananeira foi dividida em folha, pecíolo, pseudocaule, rizoma, engaço, botão floral e fruto para se analisar os macronutrientes. A partir dos dados coletados ajustou-se um programa de regressão e obteve-se as curvas de absorção de macronutrientes, cujos resultados permitiram concluir que, as plantas matriz e rebento: necessitam de expressivas quantidades de macronutrientes para desenvolver e produzir; apresentam uma absorção de macronutrientes que acompanha a matéria seca acumulada pelos órgãos, exceto para o potássio no período; absorvem mais de setenta e cinco por cento de nitrogênio, fósforo e potássio após o estádio de 180 dias e acima de setenta por cento do cálcio e magnésio, a partir de 240 e 120 dias; podem repor ao solo grande parte dos macronutrientes absorvidos; apresentam uma razão de absorção de macronutrientes de 20 K:12 N:10 Mg: 9 Ca: 1 P e de 26,5:8 N:4,5 Mg: 4,5 Ca: 1 P. Conclui-se ainda que: deve-se considerar a quantidade de macronutrientes exportados com a colheita e consumo dos frutos, principalmente nitrogênio, fósforo e potássio, em um programa de adubação; a planta rebento acumula mais macronutrientes do que a planta matriz.

Open Educational Resources : Experiences of use in a Latin-American context

This paper presents practical experiences using Open educational Resources (OER) for basic and elementary education (K12), educational research and research training on two inter-institutional projects with the collaboration of thirteen higher education institutions and with the support of the Corporación de Universidades para el Desarrollo del Internet (CUDI) and by the Consejo Nacional de Ciencia y Tecnología (CONACYT) of Mexico and hosted by the Tecnológico de Monterrey. The first initiative is titled "Knowledge Hub for K-12 Education" with the main goal of enrich a catalog of Open Educational Resources for basic and elementary education (K-12) for Mexico and Spanish speaking countries in Latin-America. The main goal of the second initiative is to build a collection of Open Educational Resources for Mobile Learning to address the issue of educational research and research training.

Effect of fertilization at sowing on nutrition and yield of crambe in second season

The interest in crambe (Crambe abyssinica ) cultivation in Brazil is on the rise, whereas information on the nutrient requirements for this crop is scarce. The objective of this work was to evaluate the effect of nitrogen-phosphorus-potassium (N-P2O5-K2O formula 8:28:16) fertilization (0, 150, and 300 kg ha-1) on crambe shoot biomass production, grain and oil yields, and nutrient extraction and exportation in the second growing season after soybean. The experiment with a Haplorthox (Dystroferric Red Latosol) was carried out for two years in Botucatu, São Paulo State, Brazil. A randomized complete block design with eight replications was used. Fertilization with NPK at sowing increased the shoot biomass production, grain yield, grain oil content, as well as nutrient extraction and exportation at harvest. In the fertilized treatments, the average amounts of nutrients extracted per hectare were 91 kg K, 71 kg N, 52 kg Ca, 9.4 kg P, 9.4 kg Mg, 7.9 kg S, 2,348 g Fe, 289 g Zn, 135 g Mn, and 18.2 g Cu; while the average values of nutrient exportation per hectare were 54 kg N, 20 kg K, 12.3 kg Ca, 10 kg P, 6.6 kg S, 3.2 kg Mg, 365 g Zn, 60 g Fe, 50 g Mn, and 7.3 g Cu, with NPK fertilizer application.

Iowa Communications Network - Fiscal Year 2012 Annual Report

The Iowa Communications Network (ICN) is the country’s premier distance learning and state government Network, committed to providing Iowans with convenient, equal access to education, government and healthcare. The Network makes it possible for Iowans, physically separated by location, to interact in an efficient, creative, and cost-effective manner. ICN provides high-speed Internet, data, video conferencing, and voice (phone) services to authorized users, under Code of Iowa, which includes: K-12 schools, higher education, hospitals, state and federal government, National Guard armories, and libraries.