Das Lektin aus <i>Pisum sativum</i>, der Gartenerbse, ist Teil der Familie der Leguminosenlektine. Diese Proteine haben untereinander eine hohe Sequenzhomologie, und die Struktur ihrer Monomere, ein all-ß-Motiv, ist hoch konserviert. Dagegen gibt es innerhalb der Familie eine große Vielfalt an unterschiedlichen Quartärstrukturen, die Gegenstand kristallographischer und theoretischer Arbeiten waren. Das Erbsenlektin ist ein dimeres Leguminosenlektin mit einer Besonderheit in seiner Struktur: Nach der Faltung in der Zelle wird aus einem Loop eine kurze Aminosäuresequenz herausgeschnitten, so dass sich in jeder Untereinheit zwei unabhängige Polypeptidketten befinden. Beide Ketten sind aber stark miteinander verschränkt und bilden eine gemeinsame strukturelle Domäne. Wie alle Lektine bindet Erbsenlektin komplexe Oligosaccharide, doch sind seine physiologische Rolle und der natürliche Ligand unbekannt. In dieser Arbeit wurden Versuche zur Entwicklung eines Funktionstests für Erbsenlektin durchgeführt und seine Faltung, Stabilität und Monomer-Dimer-Gleichgewicht charakterisiert. Um die spezifische Rolle der Prozessierung für Stabilität und Faltung zu untersuchen, wurde ein unprozessiertes Konstrukt in <i>E. coli</i> exprimiert und mit der prozessierten Form verglichen. <br /> <br /> Beide Proteine zeigen die gleiche kinetische Stabilität gegenüber chemischer Denaturierung. Sie denaturieren extrem langsam, weil nur die isolierten Untereinheiten entfalten können und das Monomer-Dimer-Gleichgewicht bei mittleren Konzentrationen an Denaturierungsmittel auf der Seite der Dimere liegt. Durch die extrem langsame Entfaltung zeigen beide Proteine eine apparente Hysterese im Gleichgewichtsübergang, und es ist nicht möglich, die thermodynamische Stabilität zu bestimmen. Die Stabilität und die Geschwindigkeit der Assoziation und Dissoziation in die prozessierten bzw. nichtprozessierten Untereinheiten sind für beide Proteine gleich. Darüber hinaus konnte gezeigt werden, dass auch unter nicht-denaturierenden Bedingungen die Untereinheiten zwischen den Dimeren ausgetauscht werden.<br /> <br /> Die Renaturierung der unprozessierten Variante ist unter stark nativen Bedingungen zu 100 % möglich. Das prozessierte Protein dagegen renaturiert nur zu etwa 50 %, und durch die Prozessierung ist die Faltung stark verlangsamt, der Faltungsprozess ist erst nach mehreren Tagen abgeschlossen. Im Laufe der Renaturierung wird ein Intermediat populiert, in dem die längere der beiden Polypeptidketten ein Homodimer mit nativähnlicher Untereinheitenkontaktfläche bildet. Der geschwindigkeitsbestimmende Schritt der Renaturierung ist die Assoziation der entfalteten kürzeren Kette mit diesem Dimer. / The lectin from <i>Pisum sativum</i> (garden pea) is a member of the family of legume lectins. These proteins share a high sequence homology, and the structure of their monomers, an all-ß-motif, is highly conserved. Their quaternary structures, however, show a great diversity which has been subject to cristallographic and theoretical studies. Pea lectin is a dimeric legume lectin with a special structural feature: After folding is completed in the cell, a short amino acid sequence is cut out of a loop, resulting in two independent polypeptide chains in each subunit. Both chains are closely intertwined and form one contiguous structural domain. Like all lectins, pea lectin binds to complex oligosaccharides, but its physiological role and its natural ligand are unknown. In this study, experiments to establish a functional assay for pea lectin have been conducted, and its folding, stability and monomer-dimer-equilibrium have been characterized. To investigate the specific role of the processing for stability and folding, an unprocessed construct was expressed in <i>E. coli</i> and compared to the processed form.<br /> <br /> Both proteins have the same kinetic stability against chemical denaturant. They denature extremely slowly, because only the isolated subunits can unfold, and the monomer-dimer-equilibrium favors the dimer at moderate concentrations of denaturant. Due to the slow unfolding, both proteins exhibit an apparent hysteresis in the denaturation transition. Therefore it has not been possible to determine their thermodynamic stability. For both proteins, the stability and the rates of association and dissociation into processed or unprocessed subunits, respectively, are equal. Furthermore it could be shown that even under non-denaturing conditions the subunits are exchanged between dimers.<br /> <br /> Renaturation of the unprocessed variants is possible under strongly native conditions with 100 % yield. The processed protein, however, can be renatured with yields of about 50 %, and its refolding is strongly decelerated. The folding process is finished only after several days. During renaturation, an intermediate is populated, in which the longer of the two polypeptide chains forms a homodimer with a native-like subunit interface. The rate limiting step of renaturation is the association of the unfolded short chain with this dimer.

Contrary to the predictions of numerous theoretical models, variation in partner quality continues to persist in mutualisms, including in the symbiosis between legumes and rhizobia. One potential explanation for the maintenance of this genetic diversity is genotype × genotype interactions, however it is unknown which genetic regions might underlie these interactions. To investigate this question, I performed a quantitative trait loci mapping experiment with two different rhizobium strains to locate potential regions of the genome influencing genotype × genotype interactions between the legume Medicago truncatula and its symbiont Sinorhizobium meliloti. I found no evidence for genotype × genotype or QTL × rhizobium interactions, however some of the QTLs colocalized with genes involved in the symbiosis signaling pathway, suggesting variation in these genes could potentially affect plant performance and fitness traits. These findings have important implications for the evolutionary interactions between legumes and rhizobia, and the genetic architecture of Medicago truncatula.

Contrary to the predictions of numerous theoretical models, variation in partner quality continues to persist in mutualisms, including in the symbiosis between legumes and rhizobia. One potential explanation for the maintenance of this genetic diversity is genotype × genotype interactions, however it is unknown which genetic regions might underlie these interactions. To investigate this question, I performed a quantitative trait loci mapping experiment with two different rhizobium strains to locate potential regions of the genome influencing genotype × genotype interactions between the legume Medicago truncatula and its symbiont Sinorhizobium meliloti. I found no evidence for genotype × genotype or QTL × rhizobium interactions, however some of the QTLs colocalized with genes involved in the symbiosis signaling pathway, suggesting variation in these genes could potentially affect plant performance and fitness traits. These findings have important implications for the evolutionary interactions between legumes and rhizobia, and the genetic architecture of Medicago truncatula.

Diverse native seed mixtures have many benefits for prairie restoration or seeded pastures. In natural grasslands, species naturally coexist with hundreds of other species in complex communities. Commercial seed mixtures rarely contain more than a small number of species, often with haphazard ratios of the component species. Thus there is no natural template for combining selected species into an optimally productive community and there is limited knowledge on how to compose a suitable species mixture. Identifying which features of a community drive increased productivity may aid in screening species and community compositions, leading to mixtures that are more specifically designed to be stable, and highly productive for the region. There is renewed interest native species as they have the potential to provide non-invasive, productive, and drought resistant rangelands that may prove more sustainable. Seven species with high agronomic potential and a broad native geographic distribution were selected for testing including: nodding brome [Bromus anomalus (Coult.)], blue bunch wheatgrass [Pseudoregneria spicata (Pursh)], western wheatgrass [Pascopyrum smithii (Rydb.)], side oats grama [Bouteloua curtipendula (Michx.)], little blue stem [Schizachyrium scoparium (Michx.)], purple prairie clover [Dalea purpurea (Vent.)], and white prairie clover [Dalea candida (Willd.)]. The early productivity and nutritional quality of these species was determined in simple mixtures in two field sites: Saskatoon and Swift Current. In the field sites the mixtures included all seven monocultures, 21 two-species mixtures and a mixture with all species. Productivity may be driven by the species richness, functional group richness, and species evenness of the community, the abundance and occurrence of particular species or functional groups, and average plant trait values within the community. Therefore, identifying the features of a community that drive increased productivity and applying them as predictive tools may aid in screening species and community compositions. Many complex mixtures of the species were planted in greenhouse experiments to determine the strongest drivers of productivity for communities of these species. The experimental approach was validated in a confirmatory experiment where optimum communities were tested. These results did not differ under a moderate drought treatment. Results were generally consistent between field and greenhouse studies. Western wheatgrass (WWG) had the highest overall plant density and the strongest effect on the forage yield of the mixtures and communities. In the field study, productivity and crude protein content were not reduced when other species were also included with WWG in the mixture. Dalea spp. did not establish as well as the other species, but had the highest crude protein concentrations. The strongest predictors of productivity were the presence and abundance of perennial C3 grasses. Increases in species richness, functional group richness, and the presence of C3s (more specifically western wheatgrass) also increased productivity, likely because of the high early relative growth rate and strong competitive ability of western wheatgrass. Overall, communities screened in the greenhouse reflected early establishment field results. The systematic approach for evaluating communities can be modified to consider enhancing other ecological functions in addition to high productivity, in other regions.

The interactions between Medicago truncatula and Phytophthora medicaginis were examined using skl, a mutant blocked in ethylene perception, and a range of wild accessions of this plant species. P. medicaginis infection of M. truncatula plants resulted in compatible responses, whereas the mutant genotype was found to be hyper-susceptible to the pathogen. Phytophthora reproduction and colonization rates of Medicago tissues supported this conclusion. Infection of skl with different pathogens reinforced this observation. Ethylene production in infected A17 and skl roots showed reduced ethylene evolution in the mutant and suggested that a positive feedback loop, known as autocatalytic ethylene production, amplified the ethylene signal. To complement the study, expression analyses of defense response genes in this interaction were studied by real time RTPCR of Phytophthora-infected and mock-infected roots. The genes analyzed were PAL, CHS, IFR, ACC oxidase, GST, and PR10. The sequences needed for the analysis were found through the scrutiny of the M. truncatula EST database employing phylogenetics and bio-informatics tools. In A17 all the genes studied were up-regulated, although the specific gene expression patterns differed. The comparison of gene expression between A17 and skl genotypes allowed the differentiation between ethylene-dependent and ethylene-independent responses. Discrete results showed that ACC oxidase homologues were downregulated in the ethylene perception mutant, corroborating the ethylene observations. However, the expression of genes involved in the phenylpropanoid metabolism was increased in skl relative to A17, suggestive of an antagonism between the ethylene perception pathway and the regulation of the phenylpropanoid pathway. This result implied that Medicago phytoalexins accumulate in the disease interaction, but raised questions about their role in resistance to Phytophthora infection. This study establishes a link between mechanisms that regulate symbiotic infection and the regulation of disease resistance to Oomycete pathogens, especially P. medicaginis. The results served to identify a series of Phytophthora-induced genes, which remain pathogen-responsive even in the absence of a functional ethylene perception pathway. While it is possible that the products of these genes are involved in resistance to P. medicaginis, the present results demonstrate that ethylene perception is required for resistance.

The journey of structural studies on lectins, starting with ConA in the 70s, has crossed many milestones. Lectins, multivalent carbohydrate-binding proteins of non-immune origin, specifically bind diverse sugar structures. They have received considerable attention in recent times on account of the realization of the importance of protein-sugar interactions, especially at the cell surface, in biological recognition. They occur in plants, animals, fungi, bacteria and viruses. Plant lectins constitute about 40% of the lectins of known structure. They can be classified into five structural groups, each characterized by a specific fold. Among them, legume lectins constitute the most extensively investigated group. Basic Winged bean lectin (WBAI) is a glycosylated, homodimeric, legume lectin with Mr 58000. The structure of WBAI complexed with methyl-a-galactose, determined earlier in this laboratory, provided information about the oligomeric state and the carbohydrate specificity of the lectin in terms of lectin-monosaccharide interactions. The present work was initiated to understand the carbohydrate specificity of the lectin, especially at the oligosaccharide level, with special reference to its blood group specificity. The hanging drop method was used for crystallizing WBAI and its complexes. Intensity data were collected on Mar Research imaging plates mounted on Rigaku RU-200 or ULTRAX-18 X-ray generators. The data were processed using DENZO and SCALEPACK of HKL suite of programs. The structure factors from the processed data were calculated using TRUNCATE of CCP4 suite of programs. The molecular replacement program AMoRe was used for structure solution. Structure refinement was carried out using the CNS software package. Model building was done using the molecular graphics program O. INSIGHT II, ALIGN, CONTACT and PROCHECK of CCP4 were used for the analysis and validation of the refined structures. WBAI exhibits differential affinity for different monosaccharide derivatives of galactose. In order to elucidate the structural basis for this differential affinity, the crystal structures of the complexes of basic winged bean lectin with galactose, 2-methoxygalactose, N-acetylgalactosamine and methyl-a-N-acetylgalactosamine have been determined. Lectin-sugar interactions involve four hydrogen bonds and a stacking interaction in all of them. In addition, a N-H O hydrogen bond involving the hydroxyl group substituted at C2 exists in the galactose and 2-methoxygalactose complexes. The additional hydrophobic interaction, involving the methyl group, in the latter leads to the higher affinity of the methyl derivative. In the lectin - N- acetylgalactosamine complex the N-H O hydrogen bond is lost, but a compensatory hydrogen bond involving the oxygen atom of the acetamido group is formed. In addition, the CH3 moiety of the acetamido group is involved in hydrophobic interactions. Consequently, the 2-methyl and the acetamido derivatives of galactose have nearly the same affinity for the lectin. The methyl group, a-linked to the galactose, takes part in additional hydrophobic interactions. Therefore, methyl-a- N-acetylgalactosamine has higher affinity than N-acetylgalactosamine to the lectin. The structures of basic winged bean lectin-sugar complexes provide a framework for examining the relative affinity of galactose and galactosamine for the lectins that bind to them. The complexes also lead to a structural explanation for the blood group specificity of basic winged bean lectin, in terms of its monosaccharide specificity. The Tn-determinant (GalNAc-a-O-Ser/Thr) is a human specific tumor associated carbohydrate antigen. Having epithelial origin, it is expressed in many carcinogenic tumors including breast, prostate, lung and pancreatic cancers. The crystal structure of WBAI in complex with GalNAc-a-O-Ser (Tn-antigen) has been elucidated, in view of its relevance to diagnosis and prognosis of various human cancers. The Gal moiety occupies the primary binding site and makes interactions similar to those found in other Gal/GalNAc specific legume lectins. The nitrogen and oxygen atoms of the acetamido group of the sugar make two hydrogen bonds with the protein atoms whereas its methyl group is stabilized by hydrophobic interactions. A water bridge formed between the terminal oxygen atoms of the serine residue of the Tn-antigen and the side chain oxygen atom of Asn128 of the lectin increase the affinity of the lectin for Tn-antigen compared to that for GalNAc. A comparison with the available structures reveals that while the interactions of the glyconic part of the antigen are conserved, the mode of stabilization of the serine residue differs and depends on the nature of the protein residues in its vicinity. The structure provides a qualitative explanation for the thermodynamic parameters of the formation of the complex of the lectin with Tn-antigen. Modelling studies indicate the possibility of an additional hydrogen bond with the lectin when the antigen is part of a glycoprotein. WBAI binds A-blood group substance with higher affinity and B-blood group substance with lesser affinity. It does not bind the O substance. The crystal structures of the lectin, complexed with A -reactive and B - reactive di and tri saccharides, have been determined. In addition, the complexes of the lectin with fucosylated A- and B-trisaccharides and with a variant of the A-trisaccharide have been modelled. These structures and models provide valuable insights into the structural basis of blood group specificities. All the four carbohydrate binding loops of the lectin contribute to the primary combining site while the loop of variable length contributes to the secondary binding site. In a significant advance to the current understanding, the interactions at the secondary binding site also contribute substantially, albeit in a subtle manner, to determine the blood group specificity. Compared to the interactions of the B- trisaccharide with the lectin, the third sugar residue of the A -reactive trisaccharide forms an additional hydrogen bond with a lysine residue in the variable loop. In the former, the formation of such a hydrogen bond is prevented by a shift in the orientation of the third sugar resulting from an internal hydrogen bond in it. The formation of this bond is also facilitated by an interaction dependent change in the rotamer conformation of the lysyl residue of the variable loop. Thus, the difference in the interactions at the secondary site is generated by coordinated movements in the ligand as well as the protein. A comparison of the crystal structure and the model of the complex involving the variant of the A-trisaccharide results in the delineation of the relative contributions of the interactions at the primary and the secondary sites in determining blood group specificity. At the disaccharide level, WBAI exhibits higher affinity for á1-3 linked Gal/GalNAc containing oligosaccharides, compared to that of other á linked oligosaccharides. With an objective of understanding the preferential binding of WBAI for á 1-3 linked Gal/GalNAc containing oligosaccharides, crystal structure of the complexes of the lectin with Galá1-4Gal, Galá1-4GalâEt and Galá1-6Gal have been determined. The reducing sugar of the disaccharides with linkages other than á1-3 binds to the lectin through a water bridge whereas the same sugar moiety with á 1-3 linkage makes direct interactions with the loop L4 of the protein. The modelling study on the complex of the lectin with Galá1-2Gal further upholds this observation. Different structures involving WBAI, reported earlier and presented here, were used to investigate the plasticity of the lectin. The front curved â-sheet, which nestles the metal binding region and on which the carbohydrate binding loops are perched, is relatively rigid. On the contrary, the flat back â-sheet, involved in the quaternary association in legume lectins, is flexible. This flexibility is probably necessary to account for the variation in quaternary structure. With the results presented in this thesis, 14 crystal structures of WBAI, in the free form and in complex with different sugars, have been reported, all from this laboratory. It is now, perhaps, appropriate to examine the new information and insights gained from these investigations, on the structure and function of the lectin. Earlier X-ray studies of WBAI contributed substantially in establishing that legume lectins are a family of proteins in which small alterations in essentially the same tertiary structure lead to large alterations in quaternary association. Structural studies on WBAI, particularly those reported here, also contributed to the elucidation of the nuances of carbohydrate recognition by lectins. A comparative study of the available structures also revealed the flexible and rigid regions of the protein. The study of the influence of covalently linked sugars on the structure of Erythrina corallodendron lectin (ECorL), a homolog of WBAI, is the content of appendix of the thesis. The three-dimensional structure of the recombinant form of Erythrina Corallodendron lectin(rECorL) complexed with lactose, has been elucidated by X-ray crystallography. Comparison of this non-glycosylated structure with that of the native glycosylated lectin reveals that the tertiary and quaternary structures are identical in the two forms, with local changes observed at one of the glycosylation sites(Asn17). These changes take place in such a way that hydrogen bonds with the neighbouring protein molecules in rECorL compensate those made by the glycan with the protein in ECorl. contrary to an earlier report, this study demonstrates that the glycan attached to the lectin does not influence the oligomeric state of the lectin. Identical interactions between the lectin and the non-covalently bound lactose in the two forms indicate, in line with earlier reports, that glycosylation does not affect the carbohydrate specificity of the lectin. The present study, the first of its kind involving a glycosylated protein with a well defined glycan and the corresponding deglycosylated form, provides insights into the structural aspects of protein glycosylation.

No description available.

Grünlandfutter mit einem hohen Ertrag und gutem Futterwert ist eine Grundvoraussetzung für die effiziente Produktion von wiederkäuenden Nutztieren. Im Zuge des prognostizierten Klimawandels werden sich die Niederschlagsmuster ändern und das Auftreten von Extremwetterereignissen, wie temporärer Trockenheit, wird sich erhöhen. Besonders produktives Grünland benötigt aber eine ausreichende und regelmäßige Wasserversorgung während der Wachstumsperiode. Deshalb werden die Futterproduktion von Grünland, die Ertragsstabilität und der Futterwert von temporärer Trockenheit beeinflusst werden. Aus diesem Grund sind Anpassungsstrategien nötig, um eine zukünftige und nachhaltige Grünlandfuttererzeugung zu sichern. Erhöhte pflanzliche Biodiversität wird oft als Möglichkeit angesehen, Funktionen von Ökosystemen, wie Produktivität und Futterwert, im Grünland zu verbessern. Es gibt eine fortlaufende Diskussion wie eine erhöhte Artenzahl auf Stress, besonders Trockenstress, reagiert und wie dabei Produktivität, Futterwert und Wassernutzung beeinflusst werden. Andere Untersuchungen zeigten, dass Artidentität und die Zusammensetzung der funktionellen Gruppen wichtige Faktoren für Produktivität und Futterwert sind. Auf Grund dessen haben wir von Juli 2009 bis Juni 2011 ein Trockenstressexperiment in einer Vegetationshalle durchgeführt. Verschiedene temporäre Trockenstressereignisse wurden in drei Aufwüchsen in zwei Vegetationsperioden durchgeführt. Die klimatischen Verhältnisse in der Vegetationshalle folgten normalen saisonalen Verläufen mit Frost im Winter und höheren Temperaturen im Sommer. Trockenstress wurde induziert, indem, nach einer anfänglichen Bewässerung, die Wasserversorgung für einen bestimmten Zeitraum eingestellt wurde. Die Wasserverfügbarkeit des Bodens konnte dabei immer kontrolliert werden. Für das Experiment wählten wir ertragsstarke und landwirtschaftlich nutzbare Arten des Grünlands der gemäßigten Zonen aus. Die Arten wurden in Monokultur und Drei- sowie Fünfartenmischungen gesät und enthielten die funktionellen Gruppen Leguminose (Trifolium repens L.), Gras (Lolium perenne L., Dactylis glomerata L.) und Kraut (Plantago lanceolata L., Taraxacum officinale F.H. Wigg. agg.). Der von uns gewählt Umfang der Artenzahl zeigte schon in anderen Biodiversitätsexperimenten einen Einfluss auf die Produktion. Untersucht wurden die Effekte von Artenzahl und funktionellen Gruppen auf Ertrag, Ertragsstabilität, Wassernutzung und Futterwert (Rohprotein, wasserlösliche Kohlenhydrate, neutrale und saure Detergenzienfasern). Als Indikatoren für die Ertragsentwicklung und die agronomische Wassernutzungseffizienz (Verhältnis von Ertrag zu Wasserverbrauch) dienten der Stickstoffertrag und die Stickstoffkonzentration der Bestände sowie δ13C Signaturen, sowohl mit unlimitierter Wasserversorgung als auch mit Trockenstress. Überdies führten wir 2009 ein Kurzzeitfeldexperiment auf einem alten Grünlandbestand auf dem Versuchsgut der Universität Göttingen in Reliehausen durch. In diesem Versuch wurde ebenso der Einfluss von Trockenstress und Artenzahl auf den Ertrag und die Wassernutzung untersucht. Unsere Daten zeigten, dass Trockenstress die Produktivität verringert und die Wassernutzung beeinflusst, beides abhängig von der Stärke des Stresses. Bei moderatem Stress war die agronomische Wassernutzungseffizienz unverändert oder stieg leicht an, bei starkem Stress verringerte sie sich jedoch. Der Stickstoffertrag und die Stickstoffkonzentration waren brauchbare Indikatoren für die agronomische Wassernutzungseffizienz, wohingegen δ13C weniger geeignet war. Die agronomische Wassernutzungseffizienz wurde von Stickstoff erhöht. Es gab keinen oder nur einen sehr geringen Einfluss von Trockenstress auf den Futterwert. Saisonale Effekte hatten mehr Einfluss auf den Futterwert. Allgemein scheint der Ertragsrückgang wichtiger als die Veränderungen des Futterwerts zu sein. Die Artenzahl beeinflusste den Futterwert und die Ertragsstabilität über die Vegetationsperiode nicht. Mit Hilfe des “sampling effect“ (Probennahmeeffekt) können der manchmal positive Einfluss der Artenzahl auf den Ertrag und die agronomische Wassernutzungseffizienz und der Rückgang dieses Einflusses unter Trockenheit erklärt werden. Mit erhöhter Artenzahl stieg der Anteil der leistungsfähigen, aber trockenheitssensitiven Leguminose. Weiterhin gab es einen Hinweis, dass die Artenzahl die Geschwindigkeit des Wasserverbrauchs erhöht. Die Ergebnisse des Feldexperiments bekräftigten die Befunde bezüglich der Effekte des Trockenstresses, des Ertrages und der Wassernutzung. Aus diesen Gründen kann die „insurance hypothesis“ (Versicherungshypothese), die besagt, dass eine erhöhte Artenzahl Ökosystemfunktionen gegenüber Umweltveränderungen stabilisieren kann, nicht bestätigt werden. Jedoch waren die funktionellen Gruppen wichtige und bestimmende Faktoren der Leistung unter nicht Wasser limitierten Bedingungen und Trockenstress. Die Leguminose hatte besonders auf Ertrag, agronomische Wassernutzungseffizienz und Rohprotein einen positiven Einfluss, jedoch erhöhte sie auch den Wasserverbrauch und die saisonale Variabilität. Gräser stabilisierten den Ertrag und den Wasserverbrauch und erhöhten die wasserlöslichen Kohlenhydrate sowie die Faserfraktionen, während sie den Ertrag und die agronomische Wassernutzungseffizienz unter den stickstofflimitierten Bedingungen unseres Experiments verringerten. Die funktionelle Gruppe Kräuter zeigte ähnliche Ergebnisse bezüglich Ertrag und Wassernutzung, aber sie erhöhte das Rohprotein. Unsere Ergebnisse demonstrieren, dass die vorhergesagte Zunahme von Trockenstressereignissen die Produktion reduzieren und die Wassernutzung ändern wird. Änderungen im Futterwert werden dabei weniger wichtig als der Ertragsrückgang sein. Für die Produktion, die Wassernutzung und den Futterwert wird die Artenanzahl weniger relevant sein als die funktionelle Zusammensetzung von Grünland. Deshalb wird eine angepasste Grasnarbenzusammensetzung für die Sicherung der Produktion von wiederkäuenden Nutztieren unter den Bedingungen des erwarteten Klimawandels Bedeutung erlangen.

The aim of the diploma thesis was to assess changes in pattern of legume storage proteins during germination. Four species of legumes were chosen for analyses ? Glycine max L., Lupinus angustifolius L., Pisum sativum L. and Vicia faba L. Seeds for analyses were sampled at the beginning, middle and end of germination. Proteins were extracted from lyophilised and homogenised material. These proteins were analysed by SDS-PAGE electrophoresis. The results proved that during seed germination the seed storage proteins cleave into smaller peptides, which forms new proteins. The intensity of protein bands in pea seeds was decreased in the area at around 48-45 kda and 40-36 kDa and the intensity of the proteins bands was increased at around the protein bands 25-23 kDa and 19-7 kDa. In lupine were not detected the protein bands over 39 kDa and during germination amount of protein bands in areas 15-7 kDa was increased

As pastagens são a base da alimentação dos ruminantes na pecuária brasileira. Sistemas de produção em condições pastoris utilizam diversos genótipos forrageiros sob diferentes formas de manejo, no intuito de otimizar respostas produtivas desses sistemas. Algumas decisões quanto ao manejo, no entanto, são tomadas de forma não criteriosa e generalista, tornando o sistema de produção frequentemente propenso a erros na gestão. A intensificação da produção animal em pastagem possibilita o aumento da participação de leguminosas forrageiras no sistema. No entanto, ainda há um limitado entendimento das respostas destas plantas quando submetidas a manejos mais intensificados. Este trabalho teve como objetivo caracterizar quatro cultivares comerciais de Arachis pintoi Krapov. & Greg., sob duas intensidades de desfolhação e dois intervalos entre desfolhas ao longo da estação de crescimento de dois anos consecutivos. Para tanto, foi conduzido um experimento analisando-se os cultivares Alqueire-1, Amarillo, Belmonte e BRS Mandobi sob quatro manejos de colheita: dois intervalos (28 e 35 dias) associados a duas intensidades de desfolha (5 e 10 cm). Determinou-se a produtividade total por estação de crescimento, acúmulo total de forragem, taxa de acúmulo diário, composição morfológica, ângulo foliar, interceptação luminosa, índice de área foliar, composição química e digestibilidade da forragem, fotossíntese foliar, fotossíntese de dossel e compostos orgânicas de reserva. O experimento foi conduzido no município de Piracicaba - SP, sendo a área experimental total de 0,11 ha. O delineamento experimental adotado foi o inteiramente casualizado com arranjo em parcelas subdivididas. À parcela foram alocados as combinações entre cultivares de A. pintoi (Alqueire-1, Amarillo, Belmonte e BRS Mandobi) e intervalos entre cortes (28 e 35 dias) em um fatorial 4 x 2. À subparcela foi designado o fator intensidade de desfolhação, na forma de duas alturas de resíduo (5 e 10 cm), tendo o experimento contado com com 3 repetições. A análise estatística dos dados foi realizada utilizando-se o procedimento MIXED do SAS&reg;. As médias dos tratamentos foram estimadas utilizando-se o LSMEANS e comparadas pelo teste Tukey a 5% de probabilidade. O cultivar Belmonte apresentou média de acúmulo total acima de 11,85 Mg MS ha-1 e taxa de acúmulo diário de 41 kg MS ha-1 nos diferentes manejos adotados, sendo o maior desempenho produtivo em relação aos demais cultivares testados. Em geral, o intervalo de 28 dias com intensidade de desfolha de 5 cm, confere maior produtividade total aos cultivares de A. pintoi. Menores produções de folíolos, pecíolos e estolões foram obtidos para o cultivar Amarillo.O perfil nutricional do cultivar Belmonte apresentou maior potencial em função da alta concentração de proteína bruta (277 g kg-1 MS) e digestibilidade in vitro da matéria orgânica (717 g kg-1 MS) em relação a Alqueire-1 e Amarillo e concentrações de fibra em detergente neutro (366 g kg-1 MS) intermediárias comparativamente aos três cultivares. Variações intraespecífica entre os cultivares apresenta pouco efeito sobre os compostos de reservas, sendo estes mais afetados pelas estratégias de manejo adotadas. Carboidratos totais não estruturais estiveram menos concentrados nos órgãos de reserva de Alqueire-1 (196 g kg-1 MO), e a concentração de N total apresentou variação somente entre os cultivares Belmonte (31 g kg-1 MO) e BRS Mandobi (25 g kg-1 MO). Em geral, a concentração de carboidratos totais não estruturais nos órgãos de reserva é maior quando o manejo de intervalos entre desfolha é de 35 dias. Plantas invasoras são mais evidentes em dosséis manejados com intensidade de desfolha de 5 cm. O cultivar Belmonte apresenta características que lhe conferem maior desempenho agronômico, sendo portanto considerado o material mais promissor. / Pastures are the backbone of the brazilian livestock industry. Pasture-based ruminant production systems make use of a diverse base of forage genotypes under various managements, aiming at the optimization of productive responses. Decisions regarding management, however, are often made under an empirical and generalistic standpoint, making these systems prone to failure. The intensification of animal production on pasture makes can be achieved with the increased participation of forage legumes in the system. However, there is still a limited understanding of the responses of these plants when subjected to more intensive management. This work aimed to characterize four commercial cultivars of Arachis pintoi (Krapov. & Greg.), under two harvest intensities and two harvest intervals throughout the summer growing season of two years. An experiment was carried out with the cultivars Alqueire-1, Amarillo, Belmonte and BRS Mandobi under four harvest management: two intervals (28 and 35 days) associated with two harvest intensities (5 and 10 cm). The total forage accumulation, daily accumulation rate, plant-part composition, leaf angle, light interception, leaf area index, chemical composition and forage digestibility, leaf photosynthesis, canopy photosynthesis and organic reserve compounds. The experiment was carried out in Piracicaba - SP. The experimental design was completely randomized with split-plot arrangement. To the plots were allocated the combinations between cultivars of A. pintoi (Alqueire-1, Amarillo, Belmonte and BRS Mandobi) and harvest intervals (28 and 35 days) in a factorial 4 x 2. To the subplot was designated harvest intensities (5 and 10 cm). Data were analyzed using the MIXED procedure of SAS&reg;. Treatment means were estimated using LSMEANS and compared by the Tukey test at 5% of probability. Belmonte had mean of total accumulation above 11.85 Mg DM ha-1 and daily accumulation rate of 41 kg DM ha-1 in the different managements adopted, the greatest production of all four entries. In general, the 28 day interval with harvest intensity of 5 cm resulted in greater total yield of all cultivars. The nutritional profile of Belmonte was the best due to the high concentration of crude protein (277 g kg-1 DM) and in vitro digestible of organic matter (717 g kg-1 DM) relative to Alqueire-1 and Amarillo and neutral detergent fiber concentrations (366 g kg-1 DM) as compared to the three cultivars. Intraspecific variations among cultivars had less effect on the reserve compounds, being more affected by the management strategies. Total non-structural carbohydrate concentratin was less in Alqueire-1 roots and stubble (196 g kg-1 OM), and total N concentration differed only between Belmonte (31 g kg-1 OM) and BRS Mandobi (25 g kg-1 OM). In general, the concentration of total non-structural carbohydrates in the reserve organs is greater when intervals between harvest is 35 days. Weeds are more evident in canopies managed with harvest intensity of 5 cm. Belmonte seems to have greater agronomic performance, being therefore considered the most promising material.