Dissertation (PhD)--University of Stellenbosch, 2007. / ENGLISH ABSTRACT: Sequence analysis showed that the essential glnA1 gene of Mycobacterium tuberculosis might be closely related to an actinomycetes progenitor sequence and that this sequence may have undergone duplication into other non-essential GS encoding genes in some Actinobateria, notably the mycobacteria. Also, the M. tuberculosis glnA1 sequence remains conserved throughout the evolution of M. tuberculosis. It was also shown that glnA1 is widely expressed in M. tuberculosis infected human pulmonary tissue, where M. tuberculosis might be present in altered phenotypes. These results imply that glnA1 is under selective pressure against evolutionary change. At transcriptional level it was shown that M. tuberculosis glnA1 might be expressed from two alternate promoter sites, but that these promoter sites may not be controlled by environmental nitrogen (in the form of ammonium) variation. We also showed that M. tuberculosis GS is effectively exported by M. smegmatis to the cell wall, but that GS secretion into the exogenous environment does not occur. Also, evidence has been presented which suggested that M. tuberculosis GS might be modified at the C-terminus, probably as part of a mechanism that retains GS in the cytosol. This hypothesis was further substantiated where it was demonstrated that two GS isoforms of different size (short isoform in cytosol, longer isoform in cell wall) are present in M. bovis BCG. It is unknown what causes this modification, since it couldn’t be observed in M. smegmatis, but it was suggested that it might be through the action of a cis- or trans-acting element present in proximity of the M. tuberculosis glnA1 gene. It was also shown that a cluster of genes found immediately downstream of the M. tuberculosis glnA1 sequence might be regulated in an operonic fashion under conditions of elevated environmental nitrogen concentrations. Two of the genes (glnE and glnA2) in this operon arrangement have been previously shown to be involved in nitrogen and glutamine metabolism. The function of the other gene, Rv2223c, is unknown. It was shown that Rv2223c homologs are mostly found in the mycobacteria and that it may encode an exported protease. It was hypothesised that this sequence and its adjacently located progenitor sequence, Rv2224c, might be involved in M. tuberculosis GS mediated metabolism. It was showed that over-expression of Rv2223c and Rv2224c may be toxic to E. coli and mycobacterial hosts, such as M. smegmatis, but that inhibition of transcription of these genes may be fatal to M. bovis BCG and M. tuberculosis H37Rv. It was also shown that Rv2223c is widely expressed in M. tuberculosis infected human tissue, which was comparable to that of glnA1. The results presented in this study shed more light on the distribution and transcriptional regulation of GS in mycobacteria and has identified new genes that might be involved in GS regulation. These results may present new approaches to tuberculosis control and thereby contribute to alleviate the burden of the disease. / AFRIKAANSE OPSOMMING: Genetiese en proteien volgorde analise het aangedui dat die glnA1 (kodeer vir glutamien sintetase (GS), ‘n essentiele protein) geen van Mycobacterium tuberculosis die naaste verwant is aan ‘n actinomycetes voorloper volgorde wat duplikasie ondergaan het om die ander nieessensiele GS koderende gene in sommige Actinobakterieë te vorm, veral in die mikobakterieë. Voords het die glnA1 geen geneties gekonserveerd gebly gedurende die evolusie van M. tuberculosis. Dit is ook aangetoon dat volop transkribasie van die glnA1 geen voorkom in die M. tuberculosis geïnfekteerde pulmonêre weefsel waar M. tuberculosis moontlik mag voorkom. Op transkriptionele vlak is dit aangetoon dat die M. tuberculosis glnA1 geen vanaf twee onderskeie promotors uitgedruk mag word, maar dat hierdie twee promotors nie deur variasies in die konsentrasie van stikstof (in die vorm van ammonium) in die omgewing beïnvloed word nie. Daar is ook aangedui dat M. tuberculosis GS effektief deur M. smegmatis oor die selmembraan na die selwand vervoer word, maar dat daar nie GS sekresie na die ekstrasellulêre omgewing geskied nie. Ook is bewyse gevind dat M. tuberculosis GS modifikasie aan die C-terminus mag ondergaan wat waarskynlik dien om GS beweging uit die sitosol te verhoed. Hierdie hipotese is versterk deurdat twee isoforms van verskillende grootte (klein in sitosol en groter in die selwand) gevind is in M. bovis BCG. Dit modifikasie meganisme is onbekend, maar vind moontlik nie plaas in nie-patogeniese mikobakterieë soos M. smegmatis nie en mag moontlik deur cis- of trans-werkende elemente gefasiliteer word. Daar is aangedui dat ‘n groepering van vier gene lanksaan die glnA1 lokus in ‘n operoniese meganisme gereguleer mag word onder variërende konsentrasies van stikstof in die omgewing. Dit is bekend dat twee van die gene in die operon (glnE en glnA2) betrokke in stikstof en gultamien metabolisme is. Die funksie van die ander twee gene (Rv2223c en Rv2224c) is onbekend. Daar is aangetoon dat volgordes soortgelyk aan Rv2223c beperk is tot die mikobakterieë en dat die geen ‘n protease, wat moontlik gesekreteer word vanuit die sitosol, kodeer. Daar is aangetoon dat die oor-produksie van die Rv2223c en Rv2224c proteine toksies is vir E. coli en mikobakterieë, soos M. smegmatis, maar dat transkripsie inhibisie hierdie gene dodelik is vir M. tuberculosis H37Rv en M. bovis BCG. Daar is ook angedui dat die ekspresie van hierdie gene volop verspreid is in M. tuberculosis geïnfekteerde menslike weefsel, soortgelyk aan diè van glnA1. Die resultate vervat in hierdie studie werp meer lig op die verspreiding en transkiptionele regulasie van GS in mikobakteriee en nuwe gene is ontdek wat betrokke by GS regulasie mag wees. Hierdie resultate mag bydra tot nuwe maniere om tuberkulose te bekamp en daardeur die voorkoms van die siekte te beperk.

Consumption of endophyte-infected tall fescue results in a syndrome of negatively altered physiological systems, collectively known as fescue toxicosis. Another challenge to endophyte-infected tall fescue -based beef cattle operations is that the soils often are selenium (Se) poor, necessitating the need to provide supplemental Se. To test the general hypothesis that different forms of supplemental Se would ameliorate the negative effects of fescue toxicosis, predominately-Angus steers (BW = 183 ± 34 kg) were randomly selected from herds of fall-calving cows grazing an endophyte-infected tall fescue pasture and consuming vitamin-mineral mixes that contained 35 ppm Se as sodium selenite (ISe), SELPLEX (OSe), or an 1:1 blend of ISe and OSe (MIX). Steers were commonly weaned and depleted of Se for 98 d. Steers were assigned (n = 8 per treatment) to the same Se-form treatments upon which they were raised and subjected to summer-long common grazing of an endophyte-infected tall fescue pasture (0.51 ppm ergot alkaloids: ergovaline plus ergovalinine; 10.1 ha). Selenium treatments were administered by daily top-dressing 85 g of vitamin-mineral mix onto 0.23 kg soyhulls, using in-pasture Calan gates. The first project objective was to determine the effect of forms of supplemental Se on whole blood Se, serum prolactin, liver glutamine synthetase (GS) activity, carcass parameters, and growth performance (Experiment 1). In Experiment 1, whole blood Se increased for all treatments from day 0 to 22 and then did not change. Across periods, MIX and OSe steers had greater whole blood Se than ISe steer. Compared to ISe steers, MIX and OSe steers had more serum prolactin. Liver GS mRNA, protein content, and activity were greater in MIX and OSe steers than ISe steers. However, the ADG and carcass parameters were not affected by Se treatments. The second project objective was to determine the effect of forms of supplemental Se on serum clinical parameters of Experiment 1 steers (Experiment 2). In Experiment 2, across periods, MIX steers had more serum albumin than OSe, and ISe steers, respectively. Serum alkaline phosphatase (ALP) activity was greater in MIX and OSe steers. In addition, blood urea nitrogen (BUN), serum sodium, phosphorus, and magnesium concentration were affected by Se treatments. Partial correlation analysis revealed that serum albumin, BUN, and ALP activity were correlated with whole blood Se concentration. The third project objective was to evaluate the hepatic transcriptome profiles of Experiment 1 steers using microarray and targeted RT-PCR analyses (Experiment 3). In Experiment 3, bioinformatic analysis of microarray data indicated that hepatic glutamate/glutamine, proline, arginine, and citrulline metabolism was affected by different forms of supplemental Se. The mRNA expression of critical proteins involved in glutamate/glutamine (GLS2, GLUD1, GLUL), proline (PYCR1, ALDH18A1), and urea (ARG1, ARG2, OAT, NAGS, OTC, ORNT1) metabolism were differentially expressed by Se treatments. Collectively, we conclude that consumption of 3 mg Se/d as OSe or MIX forms of Se in vitamin-mineral mixes 1) increased whole blood Se content, an indicator of greater whole-body Se assimilation; 2) increased serum prolactin, albumin, and ALP, the reduction of which are hallmarks of fescue toxicosis; and 3) altered hepatic nitrogen metabolism, as indicated by changes in key enzymes of glutamate/glutamine, proline, and urea metabolism. However, 4) these positive effects on metabolic parameters were not accompanied by increased growth performance.

As nitrogen (N) is a critical nutrient for plant growth, the development of synthetic N fertilisers dramatically changed agricultural production in the twentieth century. Improvement in N use efficiency (NUE) has been a focus of breeding for grain crop species, since protein is an important component of the harvested product. The study of NUE in sugarcane has lagged behind grain crops, mainly because N is not a component of sucrose, the primary product of the traditional sugarcane industry. Recently, improvement in NUE has become a focus of sugarcane breeding, due largely to environmental concerns regarding pollution from high N fertilisation, and the increasing cost of N fertilisers. This thesis aimed to gain an initial understanding of the genetic basis for variability in NUE in sugarcane. This was achieved through: (i) the screening of 168 sugarcane genotypes under limiting and non-limiting N supply in two glasshouse experiments; (ii) the mapping of marker-trait associations (MTA) for biomass and physiological traits under limiting and non-limiting N supply in a sugarcane mapping population; (iii) the analysis of expression of candidate genes encoding enzymes involved in the central processes of N assimilation and remobilisation in plants; and (iv) the mapping of candidate genes in a sugarcane genetic map. Genetic variation was identified for growth traits as well as physiological traits including %N, internal NUE (iNUE, g dry weight g-1 N) and leaf glutamine synthetase (GS) activity in a sugarcane mapping population. These traits were also analysed for linkage with genetic markers. Genetic variation in the screened genotypes was higher under limiting N supply, a finding that was reflected by the fact that marker-trait associations (MTA) for increases in iNUE were not identified under non-limiting N supply in the commercial parent of the mapping population. Contrary to findings in grain crop species, there was no link between GS activity and other traits, either through phenotypic correlations or co-location of MTA. The expression of candidate genes encoding GS, nitrate reductase (NR) and alanine amintotransferase (AlaAT) was quantified with Sequenom™ MassARRAY technology. Plants were grown under growth-limiting N supply, non-limiting N supply, or a N-pulse treatment, which consisted of growth-limiting N supply followed by non-limiting N supply 24 hours prior to sampling. Two genes, scAlaAT.d and scGS1.a, encoding AlaAT and GS respectively, were identified as non-responsive to changes in N supply, whereas scAlaAT.a, scGS1.b and scGS1.c had significantly (p<0.05) increased expression under a N-pulse, indicating an important role for these genes in the response of sugarcane to a sudden increase in N availability. The location of candidate genes associated with variation in NUE in a sugarcane genetic map were sought through restriction fragment length polymorphism (RFLP) markers. Twenty-two probes were screened, of which two generated single-dose markers, allowing the mapping of a single allele of scAspAT, encoding aspartate aminotransferase, and two alleles of scGS2, encoding plastidic GS. Because of the economic and environmental consequences of inefficient N fertiliser application, the development of sugarcane cultivars with improved NUE is essential. Since variation for NUE exists, especially in unimproved sugarcane varieties, this may be achieved through traditional breeding methods by screening existing breeding populations under limiting N supply. Additionally, an improved understanding of the genetic basis of variation for NUE in sugarcane should be pursued by further analysis of candidate gene response to changing N availability by screening widely varying cane species for differences in gene expression, enzyme activity and metabolite profiles. The further addition of candidate gene locations to sugarcane genetic maps will aid both future marker-assisted selection in breeding, and a fundamental understanding of genetic control of NUE variation. Through the development of sugarcane cultivars with improved NUE and an enhanced knowledge of the genetic control underpinning sugarcane N physiology, concerns regarding high N fertiliser applications may be mitigated and sustainability ensured.

Submitted by IVANA PAULA FERRAZ SANTOS DE BRITO null (ivanapaulaf@yahoo.com.br) on 2016-12-12T11:21:38Z No. of bitstreams: 1 Tese_Ivana Paula Ferraz.pdf: 1609743 bytes, checksum: dabfed03ce3dbb1480947a22a52cddcd (MD5) / Approved for entry into archive by Felipe Augusto Arakaki (arakaki@reitoria.unesp.br) on 2016-12-13T12:19:57Z (GMT) No. of bitstreams: 1 brito_ipfs_dr_bot.pdf: 1609743 bytes, checksum: dabfed03ce3dbb1480947a22a52cddcd (MD5) / Made available in DSpace on 2016-12-13T12:19:57Z (GMT). No. of bitstreams: 1 brito_ipfs_dr_bot.pdf: 1609743 bytes, checksum: dabfed03ce3dbb1480947a22a52cddcd (MD5) Previous issue date: 2016-11-11 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A sensibilidade de plantas daninhas a herbicidas frequentemente se apresenta variável, em função de diversos fatores. O herbicida amônio glufosinate tem ação de contato e inibe a atividade da enzima glutamina sintetase, causando dentre outros efeitos, o acúmulo de amônia nos tecidos foliares, sendo esse um bom indicativo da intoxicação das plantas. Desse modo, o objetivo desse trabalho foi identificar a resposta de plantas de Bidens pilosa e Conyza sumatrensis a diferentes doses do herbicida amônio glufosinate, e a variação da sensibilidade em populações das plantas e de suas progênies ao herbicida. Foram realizados três estudos, todos em casa-de-vegetação e repetidos em diferentes momentos. No primeiro estudo, dois experimentos de dose-resposta foram conduzidos, e foram aplicadas sete diferentes doses do herbicida amônio glufosinate (0, 50, 100, 200, 400, 800, e 1600 g ha-1), com quatro repetições, para cada espécie, sendo realizada análise de amônia e avaliações visuais de fitointoxicação. No segundo estudo, de variação de sensibilidade de B. pilosa e C. sumatrensis ao amônio glufosinate, 44 plantas de B. pilosa e 16 de C. sumatrensis foram aplicadas com a dose de 200 g ha-1 do herbicida, sendo nesse momento a região meristemática e folha mais nova de cada planta protegidas com saco plástico para que não recebessem o herbicida e possibilitasse a manutenção das plantas vivas. Dois dias após a aplicação (DAA) realizou-se a análise do teor de amônia, nas folhas expostas à aplicação, e as folhas protegidas foram mantidas nas plantas para que as mesmas se recuperassem do tratamento e produzissem sementes, utilizadas no estudo de sensibilidade de progênies. Ao atingirem o estádio reprodutivo os botões florais das plantas foram protegidos com sacos de papel para evitar a polinização cruzada e garantir a produção de sementes somente por autofecundação. No terceiro estudo, de variação da sensibilidade das progênies de B. pilosa e C. sumatrensis ao amônio glufosinate, progênies de sete das plantas anteriores foram também tratadas com 200 g ha-1 do amônio glufosinate. Foram quantificados os teores de amônia nos tecidos e porcentagens de controle foram avaliadas visualmente. A amônia foi extraída do tecido foliar fresco das plantas das duas espécies e quantificada por espectrofotometria. Foram realizadas avaliações visuais de controle aos 0, 3, 7, 14 e 21 DAA utilizando-se escala visual de notas variando de 0 a 100%. Os dados obtidos foram analisados pelo teste T e ajustados modelos de regressão não-linear. O estudo de dose-resposta demonstrou que o teor de amônia aumenta de forma assintótica com o aumento da dose do herbicida e que a porcentagem de intoxicação das espécies estudadas também aumenta de modo assintótico com o aumento tanto da dose do herbicida quanto do teor de amônia nos tecidos. O segundo estudo, de variação da sensibilidade, demonstrou haver variabilidade entre indivíduos de uma mesma população sensível ao herbicida, para as duas espécies analisadas; e o terceiro, de estudo das progênies, demonstrou que, para Bidens pilosa, as progênies das plantas com as diferentes capacidades de acumular amônia nos tecidos foram similares em termos de nível de acúmulo; no entanto, no caso de Conyza sumatrensis, as progênies de plantas com maior capacidade em acumulá-la, também exibiram maiores teores internos após o tratamento com glufosinate. Para as duas espécies, a variabilidade dos teores internos de amônia para cada progênie foi bastante alta, suplantando as diferenças médias entre as diferentes progênies, indicando pequena herdabilidade dessa característica. / The sensitivity of weeds to herbicide is often variable, due to several factors. The glufosinate-ammonium is a contact herbicide and inhibits the activity of the glutamine synthetase enzyme causing, among others, ammonia accumulation in the leaves, an indicator of the plants intoxication. The objective of this study was to evaluate the response of Bidens pilosa and Conyza sumatrensis to different doses of glufosinate ammonium and the sensitivity range of the plants and their progenies to the herbicide. Three studies were conducted, all in a greenhouse and repeated at different times. In the first study, two experiments were conducted to examine the dose-response curve, and the treatments were seven different doses of the herbicide glufosinate ammonium (0, 50, 100, 200, 400, 800, and 1600 g ha-1), with four replicates for each specie. In the second study, which examined the sensitivity range of B. pilosa and C. sumatrensis to glufosinate ammonium, 44 B. pilosa plants and 16 C. sumatrensis plants were sprayed with 200 g ha-1 of the herbicide. At the time of spraying, the meristematic region and the youngest leaf of each plant were protected with a plastic bag so that they would not receive the herbicide, thus keeping the plants alive. At two days after treatment (DAT), an analysis of the ammonium content on the sprayed leaves them was conducted. The protected leaves were kept on the plants enabling to recover from herbicide treatment and to produce seeds used to assess the sensitivity of B. pilosa and C. sumatrensis progenies to glufosinate ammonium. When the plants had reached the reproductive stage, the flower buds were covered with paper bags to prevent cross-pollination and guarantee that only self-pollination would take place. In the third study, the sensitivity range of the progeny of B. pilosa and C. sumatrensis to glufosinate ammonium was investigated; in this experiment, the progenies of seven of the previous plants were sprayed with 200 g ha-1 of glufosinate ammonium. It was measured the ammonium contents in the tissues and herbicide injury to plants was visually assessed. Ammonium was extracted from fresh leaf tissue immediately after leaf collection from the two species, and quantified per spectrophotometry. Evaluations of visual injury were conducted at 0, 3, 7, 14, and 21 DAT using a visual scale with grades ranging from 0 to 100%. The data were analyzed for t test (p≤0,05) and adjusted by non-linear regression models. The dose-response study showed that increase in ammonia content is related to the treatments used, being correlated to toxicity in the two species. The second study, the sensitivity variation showed that there was variability among individuals of the same population, for both species. The progenies study demonstrated that, Bidens pilosa, progeny plants with different capacities to accumulate ammonia in the tissues were similar in terms of buildup level; However, for Conyza sumatrensis, progeny plants with the greatest ability to accumulate it also exhibited higher internal levels after treatment with glufosinate. The variability of internal ammonia levels for each progeny was quite high, for both species, surpassing the average differences between different progenies, indicating low heritability of this characteristic.

Glutamine synthetase (GS) is a key enzyme in nitrogen (N) assimilation, especially during seed development. This thesis has identified three cytosolic GS isoforms (HvGS1) in barley (Hordeum vulgare L. cv Golden Promise). The quantitation of gene expression, isoform localisation and response to N supply has revealed that each gene plays a non-redundant role in different tissues throughout seedling development. The localisation of HvGS1_1 in vascular cells of different tissues, combined with its abundance in the stem and its response to changes in N supply, indicate that HvGS1_1 is important in N transport and remobilisation. HvGS1_1 is located on chromosome 6H at 72.54 cM, close to the marker HVM074 which is associated with a major quantitative trait locus (QTL) for grain protein content (GPC). HvGS1_1 may be a potential candidate gene to manipulate barley GPC. HvGS1_2 mRNA was localised to the leaf mesophyll cells, in both the cortex and the pericycle of roots and was the dominant HvGS1 isoform in these tissues. HvGS1_2 expression increased in the leaves with an increasing supply of N, suggesting that its role is in the primary assimilation of N. HvGS1_3 was specifically and predominantly localised in the grain, being highly expressed throughout grain development. HvGS1_3 expression increased specifically in the roots of plants grown on high NH₄⁺ suggesting that it has a primary role in grain N assimilation and also in the protection from ammonium toxicity in roots. The expression of the HvGS1 genes is directly correlated with both protein and enzymatic activity, indicating that transcriptional regulation is of prime importance in the control of GS activity in barley. Analysis of 15 different barley cultivars found no correlation between HvGS expression and various desirable attributes. Transgenics which over-express and silence individual HvGS1 isoforms have been produced and confirmed, to analyse for changes in beneficial traits.

Using a mechanism-based approach to drug discovery, efforts have been directed towards developing novel ATP mimics that can act as GS inhibitors. The purine-based systems, adenosine, adenine and allopurinol, were identified as possible scaffolds for potential ATP mimics, while various meta-disubstituted benzenoid compounds, 3-aminobenzonitrile, 3-aminophenol, resorcinol, 3-aminobenzyl alcohol, 3-hydroxybenzoic acid and 3-aminobenzoic acid have been explored as adenine analogues. These compounds were treated with different alkylating and acylating agents. Allylation of all the substrates was achieved using allyl bromide and N-9 alkylation of protected allopurinol was effected using a number of specially prepared Baylis-Hillman adducts. Acylation of the benzenoid precursors with chloroacetyl chloride, acetoxyacetyl chloride, acryloyl chloride and specially prepared 2,3,4,5,6-pentaacetylgluconoyl chloride afforded the corresponding mono- and /or diacylated products in varying yields (4-96%). Elaboration of the alkylated and acylated products has involved the reaction of hydroxy systems with diethyl chloro phosphate and chloro derivatives with triethyl phosphite in Arbuzov-type reactions to afford phosphorylated products. In all cases, products were fully characterized using 1- and 2-D NMR analysis and, where appropriate, high-resolution mass spectrometry. The application of Modgraph and ChemWindow NMR prediction programmes has been explored and the resulting data have been compared with experimental chemical shift assignments to confirm chemical structures and, in some cases, to establish the position of allylation or acylation. Experimental assignments were found to be generally comparable with the Modgraph data, but not always with the ChemWindow values. The docking of selected products in the 'active-site' of GS and their structural homology with ATP, both in their free and bound conformations have been studied using the ACCELERYS Cerius² platform. All the selected ATP mimics exhibit some form of interaction with the 'active-site' residues, and a number of them appear to be promising GS ligands.

Field and greenhouse experiments were conducted to determine barnyardgrass control as affected by glufosinate and graminicide tank-mixtures, application timing of tank-mixtures of graminicides plus glufosinate and application time of day of tank-mixtures of glufosinate and clethodim. When increased rates of graminicide were tank-mixed with glufosinate, barnyardgrass control was unaffected by quizalofop-P plus glufosinate; however, clethodim plus glufosinate control in the field indicated the potential for reduced barnyardgrass control. When evaluating increasing glufosinate rates tank-mixed with graminicides, barnyardgrass control was not negatively affected by the combination of glufosinate and graminicides. The difference in soybean yield among the graminicides may indicate that the cyclohexanedione herbicides had a slight yield advantage over the aryloxyphenoxypropionate herbicides due to potential increased levels of barnyardgrass control. Applications of glufosinate alone provide variable control throughout the growing season in both field and greenhouse experiments. Although barnyardgrass control in the field was not affected by glufosinate application timing, data from the greenhouse indicates potential exists for reduced control if glufosinate is applied 1 or 3 d before graminicides. Clethodim was unaffected by application time of day; however, glufosinate applications at midnight reduced barnyardgrass control compared to applications made at noon and 6 P.M. Applications at 6 A.M. also reduced barnyardgrass efficacy compared to applications at 6 P.M. Environmental factors such as temperature and light at the time of application are likely responsible for the time of day effects observed in these studies. For maximum benefit from incorporating graminicides into a glufosinate weed control system, fluazifop-P, quizalofop-P, clethodim and sethoxydim should be applied with glufosinate at 594 or 890 g ai ha-1. Sequential treatments of glufosinate should be applied 7 d prior to a graminicide application or 1, 3 or 7 d after a graminicide application. To optimize barnyardgrass efficacy with tank mixtures of glufosinate and clethodim, applications should be made at noon or early evening to avoid potential time of day effects.

No description available.

Deuteration is the replacement of a hydrogen atom by deuterium atom in a molecule. The replacement begins at the most acidic hydrogen in the molecule. In ATP, the deshielded hydrogen is C8-H which is the first replaced during deuteration. During ATP deuteration some of the ATP is hydrolysed to ADP concurrently. Using kinetic analysis, it was confirmed that the ATP hydrolysis that occurs is 1st order in ATP concentration, while the hydrogen replacement is 2nd order. The ATP and its C8 deuterated analogue were tested against three enzymes shikimate kinase (SK), acetate kinase (AK) and glutamine synthetase (GS) to determine if a kinetic isotope effect (KIE) exists in these systems. With AK and GS, the KIED increased as the KIEH decreased, while with SK the KIED decreased as the KIEH increased as the concentration of the ATP or deuterated analogue increased. Deuteration of imidazole and purine compounds reduced the specific activity of AK or SK at low concentrations in an enzyme-catalysed reaction. From a library of imidazole-containing compounds that inhibited SK, three compounds were selected and their IC50 values were determined on the SK-catalysed reaction. These compounds show a differential potency and efficiency between their protonated and deuterated analogues when compared in a 1:1 mixture. Synthesized purines incorporating three different substituents at N-9 were tested against AK or SK for their ability to lower the specific activity of the enzymes used / Physics / M. Sc. (Physics)

Deuteration is the replacement of a hydrogen atom by deuterium atom in a molecule. The replacement begins at the most acidic hydrogen in the molecule. In ATP, the deshielded hydrogen is C8-H which is the first replaced during deuteration. During ATP deuteration some of the ATP is hydrolysed to ADP concurrently. Using kinetic analysis, it was confirmed that the ATP hydrolysis that occurs is 1st order in ATP concentration, while the hydrogen replacement is 2nd order. The ATP and its C8 deuterated analogue were tested against three enzymes shikimate kinase (SK), acetate kinase (AK) and glutamine synthetase (GS) to determine if a kinetic isotope effect (KIE) exists in these systems. With AK and GS, the KIED increased as the KIEH decreased, while with SK the KIED decreased as the KIEH increased as the concentration of the ATP or deuterated analogue increased. Deuteration of imidazole and purine compounds reduced the specific activity of AK or SK at low concentrations in an enzyme-catalysed reaction. From a library of imidazole-containing compounds that inhibited SK, three compounds were selected and their IC50 values were determined on the SK-catalysed reaction. These compounds show a differential potency and efficiency between their protonated and deuterated analogues when compared in a 1:1 mixture. Synthesized purines incorporating three different substituents at N-9 were tested against AK or SK for their ability to lower the specific activity of the enzymes used / Physics / M. Sc. (Physics)