A MICROCOMPUTER MODEL FOR IRRIGATION SYSTEM EVALUATION

ICEASE (Irrigation Cost Estimator and System Evaluator) is a microcomputer model designed and developed to meet the need for conducting economic evaluation of adjustments to irrigation systems and management techniques to improve the use of irrigated water. ICEASE can calculate the annual operating costs for irrigation systems and has five options that can be used to economically evaluate improvements in the pumping plant or the way the irrigation system is used for crop production.Crop Production/Industries,

Structural Adaptability Facilitates Histidine Heme Ligation in a Cytochrome P450

Almost all known members of the cytochrome P450 (CYP) superfamily conserve a key cysteine residue that coordinates the heme iron. Although mutation of this residue abolishes monooxygenase activity, recent work has shown that mutation to either serine or histidine unlocks non-natural carbene- and nitrene-transfer activities. Here we present the first crystal structure of a histidine-ligated P450. The T213A/C317H variant of the thermostable CYP119 from Sulfolobus acidocaldarius maintains heme iron coordination through the introduced ligand, an interaction that is accompanied by large changes in the overall protein structure. We also find that the axial cysteine C317 may be substituted with any other amino acid without abrogating folding and heme cofactor incorporation. Several of the axial mutants display unusual spectral features, suggesting that they have active sites with unique steric and electronic properties. These novel, highly stable enzyme active sites will be fruitful starting points for investigations of non-natural P450 catalysis and mechanisms

A Panel of TrpB Biocatalysts Derived from Tryptophan Synthase through the Transfer of Mutations that Mimic Allosteric Activation

Naturally occurring enzyme homologues often display highly divergent activity with non-natural substrates. Exploiting this diversity with enzymes engineered for new or altered function, however, is laborious because the engineering must be replicated for each homologue. A small set of mutations of the tryptophan synthase β-subunit (TrpB) from Pyrococcus furiosus, which mimics the activation afforded by binding of the α-subunit, was demonstrated to have a similar activating effect in different TrpB homologues with as little as 57 % sequence identity. Kinetic and spectroscopic analyses indicate that the mutations function through the same mechanism: mimicry of α-subunit binding. From these enzymes, we identified a new TrpB catalyst that displays a remarkably broad activity profile in the synthesis of 5-substituted tryptophans. This demonstrates that allosteric activation can be recapitulated throughout a protein family to explore natural sequence diversity for desirable biocatalytic transformations

Duplex strand joining reactions catalyzed by vaccinia virus DNA polymerase

Vaccinia virus DNA polymerase catalyzes duplex-by-duplex DNA joining reactions in vitro and many features of these recombination reactions are reprised in vivo. This can explain the intimate linkage between virus replication and genetic recombination. However, it is unclear why these apparently ordinary polymerases exhibit this unusual catalytic capacity. In this study, we have used different substrates to perform a detailed investigation of the mechanism of duplex-by-duplex recombination catalyzed by vaccinia DNA polymerase. When homologous, blunt-ended linear duplex substrates are incubated with vaccinia polymerase, in the presence of Mg(2+) and dNTPs, the appearance of joint molecules is preceded by the exposure of complementary single-stranded sequences by the proofreading exonuclease. These intermediates anneal to form a population of joint molecules containing hybrid regions flanked by nicks, 1–5 nt gaps, and/or short overhangs. The products are relatively resistant to exonuclease (and polymerase) activity and thus accumulate in joining reactions. Surface plasmon resonance (SPR) measurements showed the enzyme has a relative binding affinity favoring blunt-ended duplexes over molecules bearing 3′-recessed gaps. Recombinant duplexes are the least favored ligands. These data suggest that a particular combination of otherwise ordinary enzymatic and DNA-binding properties, enable poxvirus DNA polymerases to promote duplex joining reactions

Synthesis of β-Branched Tryptophan Analogues Using an Engineered Subunit of Tryptophan Synthase

We report that l-threonine may substitute for l-serine in the β-substitution reaction of an engineered subunit of tryptophan synthase from Pyrococcus furiosus, yielding (2S,3S)-β-methyltryptophan (β-MeTrp) in a single step. The trace activity of the wild-type β-subunit on this substrate was enhanced more than 1000-fold by directed evolution. Structural and spectroscopic data indicate that this increase is correlated with stabilization of the electrophilic aminoacrylate intermediate. The engineered biocatalyst also reacts with a variety of indole analogues and thiophenol for diastereoselective C–C, C–N, and C–S bond-forming reactions. This new activity circumvents the 3-enzyme pathway that produces β-MeTrp in nature and offers a simple and expandable route to preparing derivatives of this valuable building block

Directed evolution of the tryptophan synthase β-subunit for stand-alone function recapitulates allosteric activation

Enzymes in heteromeric, allosterically regulated complexes catalyze a rich array of chemical reactions. Separating the subunits of such complexes, however, often severely attenuates their catalytic activities, because they can no longer be activated by their protein partners. We used directed evolution to explore allosteric regulation as a source of latent catalytic potential using the β-subunit of tryptophan synthase from Pyrococcus furiosus (PfTrpB). As part of its native αββα complex, TrpB efficiently produces tryptophan and tryptophan analogs; activity drops considerably when it is used as a stand-alone catalyst without the α-subunit. Kinetic, spectroscopic, and X-ray crystallographic data show that this lost activity can be recovered by mutations that reproduce the effects of complexation with the α-subunit. The engineered PfTrpB is a powerful platform for production of Trp analogs and for further directed evolution to expand substrate and reaction scope

Comportamento de gramíneas perenes recentemente introduzidas no Brasil Central

Dry matter production of fourteen recently introduced perennial grasses was compared with that of pangolagrass (Digitaria decumbens) and of "Taiwan A-24" (Digitaria pentzii) in cutting trials at the Matão Training Center of the IRI Research Institute in the State of São Paulo. Each grass was evaluated at two soil fertility levels on a red latosol. Brachiaria decumbens (IRI 562) was the outstanding grass with respect to dry matter yield over a two year period. Its yield was significantly higher than that of any of the other grasses, at both high and low fertilizer levels. Another Brachiaria species, IRI 409, and various other Digitaria species were superior to pangolagrass and to "Taiwan A-24". Additional research on animal acceptability, nutritive value, and performance under grazing is necessary for a better evaluation of this new germsplasm as promising contributions to pasture improvement in South-Central Brazil.O propósito do estudo foi avaliar gramíneas recentemente introduzidas, em comparação com o capim pangola (Digitaria decumbens) e o pangola "Taiwan A-24" (Digitaria pentzii). Foram comparadas quatorze gramíneas perenes, em um ensaio de corte, no Centro de Treinamento de Matão, do Instituto de Pesquisas IRI, no Estado de São Paulo. Cada gramínea foi avaliada em dois níveis diferentes de fertilidade de solo, em um latossolo vermelho. Brachiaria decumbens (IRI 562) foi excelente com respeito ao rendimento de matéria sêca durante um período de dois anos. Seu rendimento foi significativamente mais alto do que o de qualquer das outras gramíneas, tanto em alta fertilidade como em baixa. Uma outra Brachiaria sp., IRI 409, e várias outras Digitaria foram também superiores ao capim pangola e ao pangola "Taiwan A-24". É necessário que sejam realizados outros estudos sôbre a aceitação pelos animais, valor nutritivo e comportamento sob pastoreio, para melhor avaliação desses novos germoplasmas como contribuições promissoras para o melhoramento de pastagens no Brasil Central

Being with donkeys: insights into the valuing and wellbeing of donkeys in central Ethiopia

This paper explores the interwoven lives of donkeys and the people who depend on them for their livelihoods in central Ethiopia. Drawing on data from twelve participatory workshops, insights are elicited into the ways donkey guardians’ value and treat their donkeys. Methodologically, Participatory Rural Appraisal (PRA) and Appreciative Inquiry (AI) techniques were merged to explore the complex and multi-dimensional contributions donkeys make to participants’ lives. Findings reveal the keeping of donkeys can make the difference between destitution and modest survival, but societal perception of donkeys as low-status animals has an impact on their owners’ lives and donkey wellbeing. This research contributes empirical insights on the valuing of donkeys and enables a deepened understanding of human-donkey relations. The combination of PRA and AI in the methodology demonstrates how to formulate a compassionate and empathetic approach for exploring donkey value and wellbeing with marginalised groups

Tryptophan synthase uses an atypical mechanism to achieve substrate specificity

Tryptophan synthase (TrpS) catalyzes the final steps in the biosynthesis of L-tryptophan from L-serine (Ser) and indole-3-glycerol phosphate (IGP). We report that native TrpS can also catalyze a productive reaction with L-threonine (Thr), leading to (2S,3S)-β-methyltryptophan. Surprisingly, β-substitution occurs in vitro with a 3.4-fold higher catalytic efficiency for Ser over Thr using saturating indole, despite >82,000-fold preference for Ser in direct competition using IGP. Structural data identify a novel product binding site and kinetic experiments clarify the atypical mechanism of specificity: Thr binds efficiently but decreases the affinity for indole and disrupts the allosteric signaling that regulates the catalytic cycle