No description available.

O extrato aquoso de erva-mate, obtido a partir de folhas secas de Ilex paraguariensis, é uma bebida amplamente consumida na América do Sul. Inicialmente, nosso objetivo foi caracterizar os compostos presentes nas amostras de erva-mate disponíveis no mercado brasileiro (CH: chimarrão; T: chá mate torrado; G: chá mate torrado, comercialmente acondicionado em garrafas ou C: em copos; TS: chá mate torrado solúvel A mutagenicidade, citotoxicidade e antimutagenicidade de todas as amostras também foram avaliadas atavés do Teste de Ames na presença e na ausência de ativação metabólica. Em seguida, analisamos a amostra TS (2,5, 5,0 e 10 mg/mL) quanto a sua atividade antioxidante e antigenotóxica. Além disso, avaliamos também os efeitos da amostra TS sobre a sinalização da leptina e da insulina no hipotálamo e o estresse oxidativo hepático de ratos adultos obesos programados pela superalimentação neonatal (S). Para induzir S, o tamanho da ninhada foi reduzido a três filhotes por lactante e as ninhadas com número padrão de filhotes (dez/lactante) foram utilizadas como controle. Aos 150 dias de vida, as proles S foram subdivididas em: TS - tratados com extrato aquoso de erva-mate (1g/kg de peso corporal/dia, por gavagem) e S - recebendo água por gavagem durante 30 dias. A prole controle (C) também recebeu água nas mesmas condições do grupo S. Em nossos resultados, verificamos a presença de ácido clorogênico, cafeína e teobromina em todas as amostras analisadas. O conteúdo de compostos fenólicos nas infusões estudadas foram CH: 5,140,23; T: 4,330,01; G: 0,930,25; C: 0,800,3 e TS: 8,350,5 mg/ml. Não observamos efeito mutagênico ou citotóxico nas amostras analisadas. Um efeito antimutagênico significativo foi observado para a cepa TA97 (pré-, co- e pós-tratamento), na presença de ativação metabólica, em todas as amostras testadas. A amostra TS também apresentou um efeito antimutagênico significativo para a TA102 (pré-, co-e e pós-tratamento), na presença de ativação metabólica. Na análise exclusiva da amostra TS, observamos uma atividade antioxidante quando utilizado o ensaio de DPPH, apresentando IC50 69,3+3,1 μg/ml. Além disso, a amostra TS apresentou um efeito protetor sobre a quebra do DNA plasmidial induzida por radicais superóxido e hidroxila, de maneira dose dependente. No teste do cometa, detectamos um efeito antigenotóxico induzido pelo TS em cultura primária de células epiteliais de esôfago. Em nossos testes in vivo observamos que os animais TS não desenvolveram sobrepeso, obesidade visceral e hiperfagia. A resistência hipotalâmica à leptina não foi significativamente revertida, porém a resistência à insulina foi minimizada pelo tratamento com TS no grupo programado pela S. No fígado, TS normalizou as atividades das enzimas antioxidantes (SOD, GPx e CAT) e diminuiu os marcadores de estresse oxidativo, MDA e 4-HNE. O tratamento com TS também reduziu o conteúdo de glicogênio e triglicerídios hepáticos. Nossos resultados sugerem que a erva-mate foi capaz de proteger o DNA contra danos oxidativos, aumentou as defesas antioxidantes, melhorou a função hepática em ratos superalimentados na lactação, talvez através da modulação da sinalização hipotalâmica da insulina podendo ser, portanto, uma importante ferramenta para prevenção e tratamento de doenças relacionadas ao estresse oxidativo. / Yerba mate extract, made from dried leaves of Ilex paraguariensis, is a tea-like beverage consumed in South America. Here, we aim, firstly, to characterize the compounds present in yerba mate beverages available in the Brazilian market (chimarrao CH; mate tea T; mate tea commercially packed in bottles B and cups ready-to-drink C; and roasted yerba mate soluble tea TS). We also evaluated its mutagenic, cytotoxic and antimutagenic properties through Ames test in the presence and absence of metabolic activation. Then, we analyzed exclusively TS sample (2.5, 5.0 and 10 mg/mL) as its antioxidant activity, on plasmid DNA cleavages and upon epithelial esophagus primary culture of Wistar rats treated with a genotoxic agent. Futhermore, we evaluated the effects of mate tea (TS) upon the hypothalamic changes of leptin and insulin signaling and hepatic oxidative stress in postnatal early overfeeding (S) rats. To induce S, litter size was reduced to three pups per dam and normal litters (ten pups/dam) were used as control. In postnatal day (PN) 150, S offspring were subdivided into: S and TS groups treated, respectively, with water or mate tea (1g/kg BW/day, by gavage) during 30 days. C offspring received water. Our results showed the presence of chlorogenic acid, caffeine and theobromine in all analyzed samples. The contents of phenolic compounds in the studied infusions were CH:5,140,23; T:4,330,01; B:0,930,25; C:0,800,3 and TS:8,350,5 mg/mL. No mutagenic or cytotoxic effect was observed for all analyzed samples. A significant antimutagenic effect was observed for S.typhimurium TA97 (CoT, PreT and PosT), in the presence of metabolic activation, for all tested samples. TS sample also exhibited a significant antimutagenic effect for S.typhimurium TA102 (CoT, PreT and PosT) in the presence of metabolic activation. We did not observed an antimutagenic effect using TA98, TA100 and TA102 for CH, T, C and B samples; and no effect for TA98 and TA100 for TS sample. The exclusive analysis of TS sample, demonstrated an IC50 value of 69,3+3,1μg/ml, in DPPH radical scavenging assay. We also observed a protective effect of TS on plasmid DNA against cleavages induced by superoxide and hydroxyl radicals, in a dose dependent manner. In the comet assay, we detected an antigenotoxic effect of TS in esophagus primary cultures. Our in vivo tests showed that postnatal S rats treated with TS presented lower body weight, total body fat, total visceral fat and food intake. The hypothalamic resistance to leptin was not significantly reversed, although insulin resistance was minimized by TS in the group programmed by neonatal overfeeding. In liver, mate treatment normalized the activities of the antioxidant enzyme activities (SOD, GPx and CAT) and decreased the oxidative stress markers, MDA and 4-HNE. The treatment with TS also reduced the liver glycogen and triglycerides contents. Taken together, our results suggest that yerba mate have a potential to protect DNA against oxidative damage, increased antioxidant defences, improved liver function in adult S rats, maybe through modulation of hypothalamic insulin signaling and may be an important tool to prevent and manage oxidative stress related disorders.

Resumo: Devido às crescentes expansões demográficas e industriais observadas nas últimas décadas, o meio ambiente tem recebido uma carga significativamente crescente de efluentes domésticos, industriais e agrícolas, causando impactos severos nos ecossistemas e um potencial comprometimento à saúde humana. Dentre os efluentes domésticos, podemos citar uma gama de poluentes, tais como químicos de diversas categorias, além de contaminações por agentes biológicos diversos. Já os efluentes industriais contêm poluentes orgânicos e/ou inorgânicos, dependendo da atividade industrial. Baseando-se nestes dados, este trabalho teve como objetivo investigar, por meio de ensaios biológicos com dois organismos-teste, a possível presença de contaminantes com potencial citotóxico, genotóxico e mutagênico, que são despejados ao longo do rio Preto, inclusive na Represa Municipal de São José do Rio Preto. O material biológico utilizado neste estudo constituiu-se de sementes de Allium cepa (cebola) e peixes da espécie Oreochromis niloticus (Tilápia). Coletas de águas foram realizadas, sazonalmente, nos meses de agosto de 2006 e 2007 (estação seca) e março de 2007 e 2008 (estação chuvosa), em seis pontos distintos: Ponto 1 (P1), 8 km antes do represamento; Ponto 2 (P2), 1 km antes do represamento; Ponto 3 (P3), local de despejo do esgoto; Ponto 4 (P4), margem oposta do despejo do esgoto; Ponto 5 (P5), saída do represamento; Ponto 6 (P6), 1 km após o represamento. Análises químicas foram realizadas para todas as coletas realizadas. Para a realização do estudo, 100 sementes de Allium cepa foram submetidas à germinação, em placa de Petri, em amostras de águas coletadas nos seis diferentes pontos do rio Preto, em água ultra pura (controle negativo) e em uma substância reconhecidamente aneugênica (Trifluralina - controle positivo), sempre à temperatura ambiente... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Due to increasing population and industrial expansion observed in recent decades, the environment has received a significant increased burden of domestic industrial and agricultural sewerage, which can cause severe impacts on ecosystems, and a potential damage to human health as well. A wide range of harmful pollutants can be found in domestic effluent, such as chemicals from various categories, in addition to contamination by various biological agents. On the other hand, industrial effluents contain organic and / or inorganic pollutants, depending on industrial activity. Based on these data, this study aimed to investigate, by means of biological tests with two test-organism, the possible presence of contaminants with cytotoxic, genotoxic and mutagenic potential, which are dumped along the Preto river, an important river that flows in the region of Sao Jose do Rio Preto/SP. The biological material used in this study consisted of seeds of Allium cepa (onion) and one specie of fish (Tilapia: Oreochromis niloticus). Water samples were taken seasonally in August 2006 and 2007 (dry season) and March 2007, and 2008 (rainy season), in six distinct sites: Site 1 (S1), 8 km before the damming, Site 2 (S2), 1 km before the damming, Site 3 (S3), place of sewerage discharge; Site 4 (S4), opposite margin of sewage discharge, Site 5 (S5), end of the damming; Site 6 (S6) 1 km after damming. Chemical analyses were performed for all collected samples. For the study, 100 seeds of A. cepa were submitted to germination in Petri dishes with samples water from six different sites of the Preto river, Ultra pure water (negative control), and with an aneugenic substance (Trifluralin - positive control). For most of collection points and periods studied, root meristems cells of A. cepa, exposed to water samples collected along the Preto river, showed no significant differences... (Complete abstract click electronic access below) / Orientador: Maria Aparecida Marin-Morales / Coorientador: Maria Tecília Vilela de Azeredo-Oliveira / Banca: Regina Teresa Rosim Monteiro / Banca: Carmem Silvia Fontanetti Christofoletti / Banca: Eduardo Alves de Almeida / Banca: Mary Massumi Itoyama / Doutor

O extrato aquoso de erva-mate, obtido a partir de folhas secas de Ilex paraguariensis, é uma bebida amplamente consumida na América do Sul. Inicialmente, nosso objetivo foi caracterizar os compostos presentes nas amostras de erva-mate disponíveis no mercado brasileiro (CH: chimarrão; T: chá mate torrado; G: chá mate torrado, comercialmente acondicionado em garrafas ou C: em copos; TS: chá mate torrado solúvel A mutagenicidade, citotoxicidade e antimutagenicidade de todas as amostras também foram avaliadas atavés do Teste de Ames na presença e na ausência de ativação metabólica. Em seguida, analisamos a amostra TS (2,5, 5,0 e 10 mg/mL) quanto a sua atividade antioxidante e antigenotóxica. Além disso, avaliamos também os efeitos da amostra TS sobre a sinalização da leptina e da insulina no hipotálamo e o estresse oxidativo hepático de ratos adultos obesos programados pela superalimentação neonatal (S). Para induzir S, o tamanho da ninhada foi reduzido a três filhotes por lactante e as ninhadas com número padrão de filhotes (dez/lactante) foram utilizadas como controle. Aos 150 dias de vida, as proles S foram subdivididas em: TS - tratados com extrato aquoso de erva-mate (1g/kg de peso corporal/dia, por gavagem) e S - recebendo água por gavagem durante 30 dias. A prole controle (C) também recebeu água nas mesmas condições do grupo S. Em nossos resultados, verificamos a presença de ácido clorogênico, cafeína e teobromina em todas as amostras analisadas. O conteúdo de compostos fenólicos nas infusões estudadas foram CH: 5,140,23; T: 4,330,01; G: 0,930,25; C: 0,800,3 e TS: 8,350,5 mg/ml. Não observamos efeito mutagênico ou citotóxico nas amostras analisadas. Um efeito antimutagênico significativo foi observado para a cepa TA97 (pré-, co- e pós-tratamento), na presença de ativação metabólica, em todas as amostras testadas. A amostra TS também apresentou um efeito antimutagênico significativo para a TA102 (pré-, co-e e pós-tratamento), na presença de ativação metabólica. Na análise exclusiva da amostra TS, observamos uma atividade antioxidante quando utilizado o ensaio de DPPH, apresentando IC50 69,3+3,1 μg/ml. Além disso, a amostra TS apresentou um efeito protetor sobre a quebra do DNA plasmidial induzida por radicais superóxido e hidroxila, de maneira dose dependente. No teste do cometa, detectamos um efeito antigenotóxico induzido pelo TS em cultura primária de células epiteliais de esôfago. Em nossos testes in vivo observamos que os animais TS não desenvolveram sobrepeso, obesidade visceral e hiperfagia. A resistência hipotalâmica à leptina não foi significativamente revertida, porém a resistência à insulina foi minimizada pelo tratamento com TS no grupo programado pela S. No fígado, TS normalizou as atividades das enzimas antioxidantes (SOD, GPx e CAT) e diminuiu os marcadores de estresse oxidativo, MDA e 4-HNE. O tratamento com TS também reduziu o conteúdo de glicogênio e triglicerídios hepáticos. Nossos resultados sugerem que a erva-mate foi capaz de proteger o DNA contra danos oxidativos, aumentou as defesas antioxidantes, melhorou a função hepática em ratos superalimentados na lactação, talvez através da modulação da sinalização hipotalâmica da insulina podendo ser, portanto, uma importante ferramenta para prevenção e tratamento de doenças relacionadas ao estresse oxidativo. / Yerba mate extract, made from dried leaves of Ilex paraguariensis, is a tea-like beverage consumed in South America. Here, we aim, firstly, to characterize the compounds present in yerba mate beverages available in the Brazilian market (chimarrao CH; mate tea T; mate tea commercially packed in bottles B and cups ready-to-drink C; and roasted yerba mate soluble tea TS). We also evaluated its mutagenic, cytotoxic and antimutagenic properties through Ames test in the presence and absence of metabolic activation. Then, we analyzed exclusively TS sample (2.5, 5.0 and 10 mg/mL) as its antioxidant activity, on plasmid DNA cleavages and upon epithelial esophagus primary culture of Wistar rats treated with a genotoxic agent. Futhermore, we evaluated the effects of mate tea (TS) upon the hypothalamic changes of leptin and insulin signaling and hepatic oxidative stress in postnatal early overfeeding (S) rats. To induce S, litter size was reduced to three pups per dam and normal litters (ten pups/dam) were used as control. In postnatal day (PN) 150, S offspring were subdivided into: S and TS groups treated, respectively, with water or mate tea (1g/kg BW/day, by gavage) during 30 days. C offspring received water. Our results showed the presence of chlorogenic acid, caffeine and theobromine in all analyzed samples. The contents of phenolic compounds in the studied infusions were CH:5,140,23; T:4,330,01; B:0,930,25; C:0,800,3 and TS:8,350,5 mg/mL. No mutagenic or cytotoxic effect was observed for all analyzed samples. A significant antimutagenic effect was observed for S.typhimurium TA97 (CoT, PreT and PosT), in the presence of metabolic activation, for all tested samples. TS sample also exhibited a significant antimutagenic effect for S.typhimurium TA102 (CoT, PreT and PosT) in the presence of metabolic activation. We did not observed an antimutagenic effect using TA98, TA100 and TA102 for CH, T, C and B samples; and no effect for TA98 and TA100 for TS sample. The exclusive analysis of TS sample, demonstrated an IC50 value of 69,3+3,1μg/ml, in DPPH radical scavenging assay. We also observed a protective effect of TS on plasmid DNA against cleavages induced by superoxide and hydroxyl radicals, in a dose dependent manner. In the comet assay, we detected an antigenotoxic effect of TS in esophagus primary cultures. Our in vivo tests showed that postnatal S rats treated with TS presented lower body weight, total body fat, total visceral fat and food intake. The hypothalamic resistance to leptin was not significantly reversed, although insulin resistance was minimized by TS in the group programmed by neonatal overfeeding. In liver, mate treatment normalized the activities of the antioxidant enzyme activities (SOD, GPx and CAT) and decreased the oxidative stress markers, MDA and 4-HNE. The treatment with TS also reduced the liver glycogen and triglycerides contents. Taken together, our results suggest that yerba mate have a potential to protect DNA against oxidative damage, increased antioxidant defences, improved liver function in adult S rats, maybe through modulation of hypothalamic insulin signaling and may be an important tool to prevent and manage oxidative stress related disorders.

Cyclodextrin glucanotransferase (EC 2.4.1.19) from the alkaliphilic Bacillus sp. G-825-6 converts starch mainly to c-cyclodextrin (CD8). A combination of error-prone PCR and DNA shuffling was used to obtain variants of this enzyme with higher product specificity for CD8 and a broad pH activity range. The variant S54 with seven amino acid substitutions showed a 1.2-fold increase in CD8-synthesizing activity and the product ratio of CD7:CD8 was shifted to 1:7 compared to 1:3 of the wild-type enzyme. Nine amino acid substitutions of the cyclodextrin glucanotransferase were performed to generate the variant S35 active in a pH range 4.0–10.0. Compared to the wild-type enzyme which is inactive below pH 6.0, S35 retained 70% of its CD8-synthesizing activity at pH 4.0.

Interaktion der FO Statoruntereinheiten a und b der ATP-Synthase aus Escherichia coli Die ATP-Synthase nimmt im Energiestoffwechsel vieler Organismen eine zentrale Stellung ein und ist ubiquitär in strukturell und funktionell homologer Form bei eukaryotischen Zellen in der inneren Mitochondrienmembran, der Thylakoidmembran von Chloroplasten und in der Cytoplasmamembran von Prokaryoten zu finden. Besonders zwischen F-, V- und A-Typ ATPasen bestehen strukturelle Ähnlichkeiten im Aufbaus des Gesamtenzyms aus zwei großen Subkomplexen. Darüber hinaus weisen die F-Typ ATPasen aller Organismen hohe Sequenzhomologien auf, welche sich auch in strukturellen Gemeinsamkeiten widerspiegeln. Als "Modellenzym" dient die FOF1 ATPase aus dem Enterobakterium Escherichia coli. Es setzt sich aus acht funktionell verschiedenen Untereinheiten zusammen, die unter Hydrolysebedingungen relativ zueinander rotieren. Die Unterteilung der Enzymstruktur in Rotor (g e -c-Oligomer) und Stator (a 3b 3d ab2) erfordert das Vorhandensein einer stabilisierenden Struktur, dem sog. "second stalk". Im Hinblick auf den Mechanismus der rotierenden ATP-Synthase und dem Modell der elastischen Kopplung erscheint die Untereinheit b geeignet, um die durch das g e -c-Oligomer aufgebaute Rotationsspannung zu speichern. Wie die beiden b Untereinheiten mit den anderen FO Untereinheiten a bzw. c interagieren ist weitgehend unbekannt. In der vorliegenden Dissertation wurden die Untereinheiten a und b auf mögliche Interaktionsstellen mit anderen Enzymuntereinheiten mittels genetisch eingefügte Cysteine und anschließender chemischer Quervernetzung untersucht. In der hier vorgestellten Arbeit konnte gezeigt werden, dass es mit dem Nulllängen Cross-linker Cu(1,10-Phenanthrolin)2SO4 [CuP] in der Region bP28C-bE39C möglich ist, Quervernetzungen zur Untereinheit a zu erzeugen. Mit den heterobifunktionellen Cross-linkern Benzophenon-4-maleimid [BPM] und N-[4-(p-Azidosalicylamido)butyl]-3´-(2´-pyridydithio)propionamid [APDP] vergrößert sich diese Region. Dabei sind die a-b Interaktionen in einer gewissen Periodizität (bP28C, bL29C, bM30C, bA31C, bK38C und bE39C) zu beobachten, was für eine Beteiligung beider b Untereinheiten spricht. Neben dem immunologischen Nachweis durch Antikörper, konnte auch über ein N-terminales Polyhistidinmotiv (His12) gezeigt werden, dass eine Interaktion zwischen den Untereinheiten a und b ausbildet wird. Der aN-His12-b Cross-link kann mittels Ni-NTA Affinitätschromatographie aufgereinigt werden. b-Dimerisierungen konnten für die Reste bS60C, bL65C und die Region bY24C-bA45C nachgewiesen werden. Der relative Abstand der b Untereinheiten zueinander nimmt dabei in ihrem Verlauf vom Cytoplasma in Richtung Membran zu, wie mit den Cross-link Reagenzien CuP, BPM und APDP gezeigt werden konnte. Ausgehend von der Untereinheit a konnten für die Reste aS27C, aN33C, aA130C, aG173C, aP182C, aN184C, aS202C und aG227C ebenfalls CuP vermittelte Quervernetzungen mit der Untereinheit b nachgewiesen werden. Die Reaktion an der Position aS27C weist auf eine cytoplasmatische Lokalisation des N-Terminus hin, die in einem 6 Transmembran-Sekundärstrukturmodell vorgeschlagen wird. Mit BPM konnte die Nähe der Aminosäuren aN33C und aP182C zum c-Oligomer gezeigt werden.

In der vorliegenden Arbeit wurden die molekularen Grundlagen für die Agonistspezifität des G-Protein-gekoppelten Rezeptors GPR34 untersucht. Mittels verschiedener funktioneller Versuche konnte an ausgewählten Orthologen des Rezeptors gezeigt werden, dass, im Gegensatz zu publizierten Daten, Lysophosphatidylserin (Lyso-PS) nicht der natürliche Agonist des GPR34 ist. Lediglich an einigen cyprinoiden Subtypen des GPR34 hat Lyso-PS surrogat-agonistische Effekte. Anhand eines detaillierten evolutionären Vergleichs von Orthologen konnten Bereiche des Rezeptors ermittelt werden, welche an der Ligandenbindung, und damit an der Agonistspezifität des GPR34 beteiligt sind. Durch Übertragung dieser Bereiche vom Karpfen-GPR34-Subtyp 2a auf den humanen GPR34 konnte dieser zu einem Lyso-PS-sensitiven Rezeptor modelliert werden. Weiterhin wurde Aminoethyl-Carbamoyl-ATP (EDA-ATP) als inverser Agonist an cyprinoiden Orthologen des GPR34 identifiziert. Die Erweiterung des möglichen Ligandenspektrums von Lipiden zu Nukleotidderivaten gibt Hinweise auf die Promiskuität der Bindungsstelle des GPR34. Die Ergebnisse zeigen, dass Lyso-PS nur eine zufällige Aktivität an einigen Orthologen des GPR34 hat. Mit Identifizierung eines Nichtlipides als invers-agonistischen Liganden ist die Suche nach dem natürlichen Liganden des GPR34 noch nicht abgeschlossen und sollte auf weitere chemische Entitäten ausgeweitet werden. / Lyso-PS (lyso-phosphatidylserine) has been shown to activate the G(i/o)-protein-coupled receptor GPR34. Since in vitro and in vivo studies provided controversial results in assigning lyso-PS as the endogenous agonist for GPR34, we investigated the evolutionary conservation of agonist specificity in more detail. Except for some fish GPR34 subtypes, lyso-PS has no or very weak agonistic activity at most vertebrate GPR34 orthologues investigated. Using chimaeras we identified single positions in the second extracellular loop and the transmembrane helix 5 of carp subtype 2a that, if transferred to the human orthologue, enabled lyso-PS to activate the human GPR34. Significant improvement of agonist efficacy by changing only a few positions strongly argues against the hypothesis that nature optimized GPR34 as the receptor for lyso-PS. Phylogenetic analysis revealed several positions in some fish GPR34 orthologues which are under positive selection. These structural changes may indicate functional specification of these orthologues which can explain the species- and subtype-specific pharmacology of lyso-PS. Furthermore, we identified aminoethyl-carbamoyl ATP as an antagonist of carp GPR34, indicating ligand promiscuity with non-lipid compounds. The results of the present study suggest that lyso-PS has only a random agonistic activity at some GPR34 orthologues and the search for the endogenous agonist should consider additional chemical entities.

No description available.

No description available.

Das aus dem Apoprotein Opsin und dem kovalent gebundenen Liganden bestehende Rhodopsin dient als Modellsystem für den Aktivierungsmechanismus der größten Klasse von G-Protein-gekoppelten Rezeptoren (GPCR). Infolge einer photochemischen Reaktion vollführt Rhodopsin eine Bewegungsabfolge von Sekundärstrukturelementen, wodurch es aktiviert wird, das G-Protein bindet und den Stimulus auf zellinterne Signalwege überträgt. Mithilfe der ortsspezifischen Mutagenese wurden Mutanten des bovinen Rhodopsins erzeugt, in eine künstliche Lipidumgebung eingelagert und hauptsächlich mittels FTIR-Spektroskopie untersucht. Anhand der Y191F- und Y192F-Mutanten konnte die Translokation des transienten Gegenions der Schiffschen Base Glu181 während der Aktivierung bestimmt werden. Die Interaktionen des Tyr206 sind für die gekoppelte Bewegung von EL2 und TM5 mitbestimmend, was mittels Y206F-Mutante gezeigt wurde. Eine Anhäufung von Methioninen auf der cytoplasmatischen Seite des Rezeptors ist u.a. für das Ausklappen der TM6 zuständig. Diese Bewegung ist wichtige Determinante der Rezeptoraktivierung. Hierfür wurden insgesamt fünf Mutanten verwendet. Im zweiten, hauptsächlichen Teil der Arbeit wird das bislang kaum untersuchte humane Rhodopsin mit dem bovinen Rezeptor verglichen. Ausgehend von verschiedenen Dunkelzuständen, konnte gezeigt werden, dass die Aktivierungsmechanismen beider Rezeptoren voneinander divergieren, um letztlich bei der Bildung der aktiven Spezies wieder zu konvergieren. Über die Analyse der Aminosäuresequenzen der Mammalia-Rhodopsine wurden zwei Bereiche hoher Variabilität identifiziert, die u.a. die molekulare Ursache für diese Diskrepanzen liefern. Diese Feststellung wurde mit human-bovinen-Rhodopsinchimären bewiesen. Ergänzend zu dieser Studie wurde Schafsrhodopsin einem Vergleich sowohl mit bovinem als auch mit humanem Rezeptor unterzogen. Es zeigte, als eine weitere natürlich vorkommende Variante des Lichtrezeptors, einen eigenständigen Weg der Aktivierung. / Rhodopsin, which consists of the apoprotein opsin and its covalently bound ligand, is used as a model system to understand the activation mechanism of the large family of G protein coupled receptors (GPCRs). As a result of a photochemical reaction, rhodopsin undergoes activating structural changes, enabling it to bind the G protein and transmitting the stimulus to intracellular signaling pathways. In the first part of this work, site-directed mutants of bovine rhodopsin were produced, incorporated into an artificial lipid environment, and studied mainly by FTIR spectroscopy. The translocation of the transient Schiff base counterion (Glu181) during the activation process was determined using the Y191F- and Y192F-mutants. The interactions of Tyr206 contributed to the coupled movement of EL2 and TM5, which was shown by Y206F-mutant. A striking accumulation of methionines on the cytoplasmic side of the receptor was observed to be a key-player for the activating outward motion of TM6. In the second and primary part of this work, human rhodopsin, which has been rarely studied, was compared with the bovine receptor. Starting from various dark states, it was shown that the activation mechanisms of both receptors diverge from each other and yet ultimately converge in the formation of the active species. By analyzing the amino acid sequences of mammalian rhodopsins, two regions of high variability were identified, which provide the molecular basis for these discrepancies. This finding was verified by the investigation of human/bovine rhodopsin chimeras. In addition to this study, ovine rhodopsin was compared with both the bovine and human forms. It showed, as another naturally occurring variant of the light receptor, an independent pathway of activation.