The study was to investigated the gene expression of light harvesting complex II (LhcII) of photosystem II (PSII) and Photosystem I reaction center subunit III (psaF) in the marine macroalga Ulva fasciata Delile in relation to copper (50 £gM CuSO4) and hypersalinity (90‰ ASW) stress. Excess copper had little effect on photosynthetic ability and gene expression of LhcII and psaF. Hypersaline decreased Fv/Fm and Fv'/Fm', but increased LhcII and psaF transcript level to a plateau after 6 h. The effects of ROS scavengers(5 mM dimethylthiourea (DMTU)¡B10 mM 4,5-dihydroxy-1,3-benzenedisulfonic acid (Tiron) or 10 mM sodium benzoate (SB) to 90‰-grown thalli) on LhcII and psaF gene expression were examined by exogenous application of. Indicate that O2•¡Ð is the factor for mediating LhcII gene expression by 90‰ while O2•¡Ð¡BH2O2 and OH• are related to the induction of psaF gene expression by hypersalinity. Exogenously applied spermidine (Spd) and spermine (Spm) recovered Fv/Fm and Fv'/Fm' and decreased LhcII and psaF transcript levels by 90‰ while putrescine (Put) recovered only Fv'/Fm' but not effect on Fv/Fm and LhcII transcript levels were increased. Electron-transport inhibitors, 50 £gM 3-(3,4 dichlorophenyl) -1,1-dimethyl urea (DCMU) or 50 £gM 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB) , applied to the 30‰-grown thalli decreased Fv/Fm and Fv'/Fm' but increased LhcII transcript levels while psaF transcript levels were decreased.It could be concluded that the expression of LhcII and psaF were influenced by the redox state of plastoquinone pool (PQ pool) and photosynthetic electron flow ; Reactive oxygen species produced under hypersaline condition influenced the LhcII and psaF transcript level, and polyamine might play a protective role against salt stress by the modulation of LhcII and psaF gene expression.

Photosynthesis in higher plants relies upon collection of light by chlorophyll molecules associated with light harvesting chlorophyll a/b-binding (LHC) proteins. The two most abundant of these are Lhcb1 and Lhcb2, which make up light harvesting complex (LHC) II trimers. They are also involved in facilitating state transitions, a process during which energy balancing between photosystem (PS) II and I is achieved. Overexcitation of PSII reduces the plastoquinone pool which activates STN7, a kinase, that phosphorylates a threonine residue on Lhcb1 and Lhcb2. In order to studythe kinetics of this we developed antibodies capable of recognizingphosphorylated forms of each of these proteins. This showed that Lhcb2 is more rapidly phosphorylated than Lhcb1, that there are no differences in the migration of phosphorylated and non-phosphorylated forms of Lhcb1 and Lhcb2 and that the majority of phosphorylated LHCII (P-Lhcb1 and PLhcb2) are associated with super- and megacomplexes. Furthermore, a state 2-specific LHCII-PSI-LHCI band contains P-Lhcb2 but almost no P-Lhcb1, and a band corresponding to M trimers (band 4 from sucrose gradients, composed of LHCII, CP24 and CP29), contains only P-Lhcb1 but no P-Lhcb2. We also developed artificial microRNA lines specifically depleted in either Lhcb1 or Lhcb2, amiLhcb1 and amiLhcb2 respectively. We show that the roles of Lhcb1 and Lhcb2 in state transitions are complementary. Lhcb1 modulate the size of grana stacks. In the absence of Lhcb1 only a few LHC trimers are formed, while in the absence of Lhcb2, the antenna looks like in the wild type although the plants cannot perform state transitions normally. Trimers containing P-Lhcb2 functionally detach from PSII and connect to PSI to balance the relative excitation pressure. State transitions only occur when both Lhcb1 and Lhcb2 are present, presumably in a (Lhcb1)2 Lhcb2 heterotrimer. In absence of Lhcb2, the LHCII-PSI-LHCI supercomplex is not formed indicting that P-Lhcb2 mediates attachment of LHCII to PSI. We tried complementing amiLhcb2 with modified Lhcb2 genes coding for proteins with altered amino acids, Arg2 to Lys or the phosphorylatable Thr3 residue to Asn or Ser. Introduction of the additional gene often causes loss of amiRNA-inhibition, however we could confirm that substitution of the Thr3 with Asn led to the absence of Lhcb2 phosphorylation and thus no state transition. / Klorofyll a/b-bindande proteiner (s k light harvesting chlorophyll a/b-binding proteins eller LHC proteiner) är viktiga för högre växters fotosyntes, då deras klorofyllmolekyler skördar solljuset. Två av dessa proteiner, Lhcb1 och Lhcb2, bygger upp ”LHCII trimerer” och finns i större mängd än de andra och dessa är även viktiga för s k ”state transtions”, en process som ser till att fotosystem (PS) I och PSII exciteras lika mycket. Om PSII exciteras för mycket reduceras plastoquinon-poolen som i sin tur aktiverar ett proteinkinas, STN7, som fosforylerar en av Lhcb1/Lhcb2s treoniner. För att studera denna fosforylering har vi utvecklat antikroppar som är specifika för dessa fosforylerade former av proteinerna, och vi använde dem för att visa att Lhcb2 fosforyleras snabbare än Lhcb1, och att största delen av det fosforylerade proteinerna (P-Lhcb1 och P-Lhcb2) finns i s k super- eller megakomplex. Ett komplex som bara finns finns i ”state 2” består av LHCII, PSI och LHCI, och det innehåller endast P-Lhcb2 men nästan inget P-Lhcb1, och ett band som består av LHCII, CP24 och CP29 innehåller endast PLhcb1. Vi skapade artificiella mikro-RNA-linjer, amiLhcb1 och amiLhcb2, som saknade antingen Lhcb1 eller Lhcb2. Lhcb1 påverkar höjden av grana stackarna. Med hjälp av dessa visade vi att Lhcb1 och Lhcb2 har komplementära roller för state transitions, saknas Lhcb1 gör växten bara få LHCII trimerer, medan om Lhcb2 gör växten antennener som liknar vildtypens, men den kan inte utföra state transitions som den. Mängden Lhcb1 påverkar storleken av ”grana stacks”. Trimerer som innehåller PLhcb2 kopplas över från PSII till PSI för att balansera excitationstrycket. Både Lhcb1 och Lhcb2, antagligen i trimerer bestående av en Lhcb2 och the Lhcb1, behövs för state transitions. Saknas Lhcb2 bildas inga komplex bestående av LHCII, PSI och LHCI, vilket visar att P-Lhcb2 antagligen möjliggör LHCIIs bindning till PSI. Vi försökte komplementera amiLhcb2 med Lhcb2 gener där amino syror bytts ut, Arg2 till Lys eller den fosforylerbara Thr3 till Asn eller Ser. När denna gen introducerades försvann dock ofta amiRNA-inhiberingen, men vi kunde visa att om Thr3 ersattes med Asn skedde inga state transitions.

This thesis contains quantum chemical models and force field calculations for the RuBisCO isotope effect, the spectral characteristics of the blue-light sensor BLUF and the light harvesting complex II. The work focuses on the influence of the environment on the corresponding systems. For RuBisCO, it was found that the isotopic effect is almost unaffected by the environment. In case of the BLUF domain, an amino acid was found to be important for the UV/vis spectrum, but unaccounted for in experiments so far (Ser41). The residue was shown to be highly mobile and with a systematic influence on the spectral shift of the BLUF domain chromophore (flavin). Finally, for LHCII it was found that small changes in the geometry of a Chlorophyll b/Violaxanthin chromophore pair can have strong influences regarding the light harvesting mechanism. Especially here it was seen that the proper description of the environment can be critical. In conclusion, the environment was observed to be of often unexpected importance for the molecular properties, and it seems not possible to give a reliable estimate on the changes created by the presence of the environment. / Diese Arbeit beinhaltet quantenchemische und molekularmechanische Modelle zum Isotopeneffekt des Enzyms RuBisCO, der spektralen Charakterisierung des Blaulicht-Rezeptors BLUF und dem Lichtsammelkomplex II (LHCII). Es wurden vor allem die Einflüsse der Umgebung auf die entsprechenden Systeme untersucht. Für RuBisCO wurde gefunden, dass der Isotopeneffekt nur marginal von der Umgebung abhängt. Im Falle der BLUF Domäne wurde eine Aminosäure charakterisiert (Ser41), die bis dato experimentell noch nicht beschrieben war. Es wurde festgestellt, dass Ser41 hochmobil ist und einen systematischen Einfluss auf die spektrale Verschiebung des BLUF Chromophors (Flavin) hat. Schließlich wurde bei LHCII festgestellt, dass kleine Veränderungen in der Geometrie eines Chlorophyll b/Violaxanthin Chromophorenpaares bereits massive Einflüsse auf den Mechanismus des Lichtsammelprozesses haben können. Insbesondere hier zeigt sich, wie kritisch die genaue Beschreibung der Umgebung ist. Zusammenfassend wurde beobachtet, dass sich die Umgebung in oft unerwarteter Weise auf die molekularen Eigenschaften auswirken kann und es daher nicht möglich zu sein scheint, die entsprechenden Effekte vorher abzuschätzen.

La photosynthèse oxygénique fournit l’énergie pour presque toutes les formes de vie sur Terre. Les premières étapes sont réalisées par quatre complexes membranaires. Deux sont des photosystèmes (PS) divisés en un core et une antenne qui, chez les plantes, est composée des protéines Lhca et Lhcb respectivement pour le PSI et le PSII. Les Lhcb les plus abondantes forment les hétérotrimères LHCII. Pour optimiser leur croissance, les organismes photosynthétiques doivent réguler précisément l’absorption de la lumière. Dans cette thèse, nous nous sommes focalisés sur un de ces mécanismes chez Arabidopsis thaliana : les transitions d’état. Cette régulation intervient en lumière faible ou fluctuante pour répondre à un déséquilibre d’excitation des photosystèmes. Elle implique la phosphorylation de Lhcb1 et Lhcb2, deux des isoformes composant le LHCII, et leur déplacement du PSII au core du PSI où elles agissent comme antenne. Récemment, l’attachement d’un LHCII additionnel à l’antenne du PSI a été rapporté, sans indice sur le rôle de la phosphorylation dans la liaison. Nous nous sommes attachés à quantifier la phosphorylation du LHCII dans les supercomplexes PSI et PSII et à déterminer les rôles respectifs de Lhcb1 et Lhcb2 dans les transitions d’état. Nous avons établi que la phosphorylation d’un seul Lhcb2 est suffisante pour l’attachement d’un LHCII au core du PSI. Nous avons ensuite isolé un complexe PSI-LHCII2 et déterminé que la liaison du second LHCII implique aussi la phosphorylation d’un seul Lhcb2. Ce travail apporte de nouvelles preuves des rôles divergents des isoformes Lhcb dans la régulation de la photosynthèse, qui sont discutées ici à la lumière de leur évolution. / Oxygenic photosynthesis directly or indirectly provides energy for almost all forms of life on earth. The first steps are performed by four membrane complexes. Two of them are photosystems (PS) organized in a core complex and an antenna system, which is composed in green organisms by Lhca and Lhcb proteins for PSI and PSII, respectively. The most abundant Lhcb proteins form the LHCII heterotrimers. To optimize growth, photosynthetic organisms have to precisely regulate their light harvesting. In this thesis, we focused on one of these mechanisms in Arabidopsis thaliana: state transitions. This regulation occurs in low or fluctuating light to answer for the imbalance of excitation between photosystems. It involves the phosphorylation of Lhcb1 and Lhcb2, two of the isoforms composing LHCII. The phosphorylated LHCII trimers detach from PSII and bind to PSI core and act as an antenna for it. Recently the attachment of additional LHCII trimers to PSI on its antenna side has been reported, with no clue of the role of phosphorylation in this binding. We set on quantifying the LHCII phosphorylation of purified PSI and PSII supercomplexes to determine the respective roles of the Lhcb1 and Lhcb2 isoforms in state transitions. We established that the phosphorylation of a single Lhcb2 protein is sufficient for the binding of PSI core. We then isolated a PSI-LHCII2 supercomplex, and determined that the binding of the additional trimer also involves the phosphorylation of a single Lhcb2 isoform per trimer. This work brings new evidences for the divergent roles of the Lhcb isoforms in light harvesting and its regulation, which are discussed here in the light of their evolution.

No description available.

Das Anliegen dieser Arbeit besteht darin, die Kinetik von elektronischen Singulett- und Triplett-Anregungen innerhalb des solubilisierten Lichtsammelkomplexes des Photosystem II (LHCII) zu untersuchen. Die Untersuchungen gliedern sich in drei Teilkomplexe: Klärung der Frage, in welchem Maße die gegenseitige Wechselwirkung zwischen angeregten Singulett-Zuständen (Singuletts)zu deren Vernichtung führt. Um dies zu analysieren, wurden Messungen zur nichtlinearen Fluoreszenz von LHCII in einem großen Bereich der Anregungsimpuls-Intensität durchgeführt. Dazu war es notwendig, eine Methode zu entwickeln, die es erlaubt, die von der Probe ausgesandte Fluoreszenz räumlich zu selektieren. Die so gemessene Fluoreszenzausbeute von solubilisiertem LHCII verringert sich um bis zu 4 Größenordnungen bei maximaler Intensität und entspricht einem sättigenden Verhalten der Fluoreszenz. Aus diesen Untersuchungen ergibt sich, daß der Transfer der Anregungsenergie innerhalb eines gesamten solubilisierten LHCII-Trimers extrem schnell vonstatten geht. Untersuchung zur Kinetik der Karotinoid-Triplett-Bildung im LHCII. Dazu wurde ein Meßaufbau für Pump-Test-Messungen der nichtlinearen Transmission mit optischer Zeitverzögerung zwischen Pump- und Test-Impuls realisiert. Besonderes Augenmerk wurde auf die Vermeidung von Meß-Artefakten gelegt. Die zeitliche Änderung der Absorption bei 507 nm spiegelt die Kinetik der Karotinoid-Triplett-Bildung in solubilisiertem LHCII wider. Bei der Modellierung der gemessenen Daten wurde die Rate für den Triplett-Transfer von 3Chl nach Car variiert. Die beste Anpassung ergibt den Grenzfall . Als untere Grenze kann im Rahmen der Meßgenauigkeit ein Wert von (0.5 ns)-1 angegeben werden. Dieser Wert ist mehr als eine Größenordnung größer, als der noch vor kurzer Zeit akzeptierte Wert von Kramer und Mathis (1980). Dieses Ergebnis zeigt, daß die Wechselwirkung zwischen Chlorophyllen und Karotinoiden in LHCII wesentlich stärker ist, als bisher angenommen wurde. Untersuchung der Wechselwirkung von Singuletts mit langlebigen Tripletts anhand der Fluoreszenz, die durch einen elektronisch verzögerten Test-Laser-Impuls hervorgerufen wird. Diese "Test-Fluoreszenz" nimmt mit Erhöhung der Pump-Intensität drastisch ab und erreicht bei hohen Pump-Intensitäten wenige Prozent des Ausgangswertes. Das Zurückkehren zum Ausgangswert wurde als Funktion der Verzögerungszeit bei verschiedenen Pump-Intensitäten gemessen. Mit Hilfe der Stern-Volmer Gleichung lassen sich daraus die quenchenden Populationen berechnen. Dies ergibt, daß das Quenching bei vergleichsweise geringen Pump-Intensitäten eindeutig den Karotinoid-Tripletts zugeordnet werden kann. Bei hohen Pump-Impuls-Intensitäten wurde eine zusätzliche, extrem stark quenchende Spezies identifiziert, bei der es sich möglicherweise um Chlorophyll-Tripletts oder -Ionen handelt. / In this study the kinetics of electronically excited singlet and triplet states within solubilized light harvesting complexes of photosystem II (LHCII) is investigated. It is subdivided into three parts: The mutual interaction between excited singlet states and their annihilation was analyzed. For that purpose nonlinear fluorescence measurements were performed in a huge range of excitation pulse intensity. A method was developed that allows a spatial selection of the fluorescence. The fluorescence yield determined in this way shows a drastic decrease of up to 4 orders of magnitude at the highest intensity used and corresponds to a saturation like behavior of the fluorescence. As a result of the theoretical investigation of this result, the excitation energy transfer within the overall LHCII-trimer is extremely fast. The kinetics of carotenoid triplet formation in LHCII was analyzed by pump-probe measurements of nonlinear transmission using an optically generated delay between pump- and probe-pulses. Special care was taken to prevent artifacts in this kind of measurement. The temporal change of the absorbance at 507 nm reflects the kinetic of carotenoid triplet formation in solubilized LHCII. The fit of the experimental data by a kinetic model provides that the rate for the triplet transfer from 3Chl to Car is (0.5 ns)-1. This value is more than one order of magnitude higher than the results obtained so far (Kramer and Mathis, 1980). This shows that the interaction between chlorophylls and carotenoids is much higher than previously assumed. The interaction between singlets and (long living) triplets was investigated by measurements of the fluorescence originating from a delayed probe pulse in the presence of a pump pulse. This "probe-fluorescence" decreases drastically with increasing pump pulse intensity up to a level of a few percent at the highest intensity. The recovery to the initial value (pump pulse off) was studied as a function of the electronically generated time delay and at different pump intensities. The population of quenching species was calculated by means of the Stern-Volmer Equation. As a result of this analysis the quenching at low pump intensities can clearly be attributed to carotenoid triplets. At high pump pulse intensities an extremely strong quenching species is formed which can be probably identified by chlorophyll-triplets or -ions.

Les caroténoïdes jouent un rôle essentiel dans les premiers événements photosynthétiques. Ils absorbent la lumière et transfèrent l'énergie résultante en excitation aux molécules voisines, assurant le respect de la succession des étapes photosynthétiques. En plus de l'absorption de la lumière, les caroténoïdes protègent l'appareil photosynthétique du stress photo-oxydatif survenant en condition de lumière intense, évitant les éventuels dommages. Les propriétés électroniques des caroténoïdes sont à la base de leurs mécanismes d'action et dans ce travail de recherche une combinaison de techniques biochimiques et spectroscopiques est utilisée pour examiner plus loin ces mécanismes avec un accent mis sur le rôle photoprotecteur joué par des caroténoïdes. Les d'échantillons analysés représentent différent niveaux d'organisation des protéines collectrices de lumière contenants ces pigments. Dans cette thèse quatre études principaux ont étés réalisés pour comprendre comment: les propriétés d'absorption des caroténoïdes lutéine et -carotène près les plantes peuvent être réglées in vivo par le site de liaison à leur protéines, le majeur complexe de capture de la lumière (LHCII) et le photosystème II (PSII) respectivement; l'altération des gènes de la voie biosynthétique des caroténoïdes peut indirectement provoquer une altération du transport d'électrons dans l'organisme photosynthétique; des molécules artificielles sont capable d'imiter le mécanisme photoprotecteur de transfert d'énergie entre les états de triplet des chromophores en mimant les protéines de l'apparat photosynthétique; la flexibilité structurelle de l'LHCII peut être explorée en modifiant son environnement. / Carotenoids play an essential role in the first steps of photosynthesis. They absorb light and they transfer the resulting excitation energy to the neighboring molecules, guaranteeing the correct order of the photosynthetic events. Additionally, carotenoids are able to protect the photosynthetic apparatus from the oxidative stress occurring in high light condition. Biological functions of carotenoids involving interaction with light, such as photosynthesis, are determined by the electronic properties of the conjugated polyene chain that is characteristic of carotenoid molecules. Understanding how these properties are tuned, is essential for understanding the mechanisms underlying carotenoid functions. Here we show that, by using a combination of different spectroscopic and biochemical approaches, these characteristics can be assessed in different kind of samples having the carotenoid molecules as common denominator. In this thesis four major studied have been performed in order to study how: the absorption properties of the two -carotenes molecules in PSII-RC and those of the two luteins in LHCII are tuned in vivo by their protein binding site, the alteration of the genes involved in the biosynthetic pathway of carotenoids has a pleiotropic effect on the photosynthetic organisms, artificial constructs are able to reproduce the photoprotective mechanism of triplet-triplet energy transfer between chromophores by mimicking the naturally occurring photosynthetic proteins, the structural flexibility of the major light harvesting complex can be probed by modifying its surrounding environment.

Oxygenic photosynthesis is the major producer of both oxygen and organic compounds on earth and takes place in plants, green algae and cyanobacteria. The thylakoid membranes are the site of the photosynthetic light reactions that involve the concerted action of four major protein complexes known as photosystem II (PSII), cytochrome b6f complex, ATP synthase and photosystem I (PSI). The function of PSII is of particular interest as it performs the light–driven water splitting reaction driving the photosynthetic electron transport. My thesis addressed different aspects of PSII assembly and the functions of its low molecular weight PSII subunits PsbX and PsbW. Photosynthesis in green algae and higher plants is controlled by the nucleus. Many proteins of nuclear origin participate in the regulation of the efficient assembly of the photosynthetic protein complexes. In this investigation we have identified one of these nuclear encoded auxiliary proteins of photosystem II, REP27, which participates in the assembly of the D1 reaction center protein and repair of photodamaged PSII in the green algae Chlamydomonas reinhardtii. Interestingly, PSII is specially enriched in Low Molecular Weight (LMW) subunits that have masses less than 10kDa. These proteins account for more than the half of the PSII subunits. Several questions remains poorly understood regarding the LMW: Which is their evolutionary origin? What function do they perform in the protein complex? Where are they located in the protein structure? In this investigation the functions of two of these LMW subunits (PsbX and PsbW) have been studied using antisense inhibition and T-DNA knockout mutant plants in Arabidopsis thaliana. Deficiency of the PsbX protein leads to impaired accumulation and functionality of PSII. Characterization of PsbW knock-out plants show that PsbW participates in stabilization of the macro-organization of PSII and the peripheral antenna (Light Harvesting Complex, LHCII) in the grana stacks of the chloroplast, also known as PSII-LHCII supercomplexes.

Study of the Arabidopsis thaliana knockout mutant lacking Lhcb3 (koLhcb3) have revealed a close similarity to the wild type plants. Growth rate, NPQ, qP, Φ(PSII), circular dichroism spectra, pigment composition and content of LCHII trimers have been found to be unaffected by this mutation. The proteomic analysis shows only some minor increases in the amount of Lhcb1 and Lhcb2. PAM fluorometry revealed a significant increase in the rate of the state 1 to state 2 state transition in the koLhcb3. None the less, the extent of state transition is identical to wild type. Alterations in the PSII-LHCII supercomplex structure have been demonstrated as well. The M-trimer was found to be rotated ~21° CCW. This altered binding of the LHCII M-trimer is likely the cause of the altered affinity resulting in the increased rate of state transition. Proteomic analysis of the phosphorylation of LHCII revealed a significant increase in state 1 and 2 LHCII phosphorylation relative to wild type. Investigation whether phosphorylation or the altered LHCII binding is the cause of the accelerated rate of state transition have not been conclusive so far. A Lhcb6 depleted mutant (koLhcb6) showed a significant alteration of the PSII-LHCII supercomplex structure and photosynthetic acclimation processes. The LHCII M-trimer is depleted in the PSII-LHCII supercomplexes causing the state transition process to be “stuck” in state 2 and the mutants ability to preform NPQ is inhibited as well. The Lhcb6 protein was concluded to be essential for the binding of the LHCII M-trimer to the PSII core as well as energy transfer. The depletion of LHCII M-trimer was linked to the reduced ability to photoacclimate using NPQ as well.

Global energy demands have escalated over the past few decades, creating a necessity for alternative energy sources. Solar technologies inspired by the primary solar energy storing process known on earth, photosynthesis, have subsequently gained popularity. The natural photosynthetic apparatus comprises a network of membrane-bound pigment-protein complexes, with the main plant light-harvesting complex (LHCII) consisting of chlorophyll (Chl) and carotenoid (Car) pigments. Electronic excitation energy transfer (ET) of the harvested energy takes place amongst these pigments on ultrafast timescales. This energy is funnelled towards a photosynthetic reaction centre where charge separation is achieved, creating a Biobattery, which powers the subsequent manufacture of energy-rich chemical compounds for photosynthetic activity. Transient absorption pump-probe spectroscopy has proven to be a useful technique for monitoring the evolution of the excited state dynamics, such as electronic transitions and excitation ET amongst Car and Chl pigments of LHCII trimers isolated from spinach leaves. This method was utilized to probe samples excited under four different conditions: at pump excitation wavelengths (𝜆𝑒𝑥) of 489 nm (preferentially exciting Cars Lutein1 and Neoxanthin) and 506 nm (targeting Cars Lutein2 and Violaxanthin), each with an intensity of either 800 nJ/pulse (relatively high) or 500 nJ/pulse (comparatively low). A global analysis was applied to each dataset using the robust, open-source Glotaran software, from which three kinetic decay lifetimes for the various processes were extracted. General spectral observations encompassed a negative pump ground state bleach (GSB) at each 𝜆𝑒𝑥; negative Chl b and Chl a GSBs, superimposed with negative stimulated emission (SE) signals; and a positive excited state absorption (ESA) band. The first lifetime of a few picoseconds corresponded mainly to Car-S2 depopulation, resulting either from energy relaxation towards Car-S1, or ET to Chls. Small, but distinct Chl b signals of less than 3 mOD were also detected on this timescale. The second lifetime, which is between 10 and 12 ps, was characteristic to the Lutein Car-S1 lifetime, mainly depicting Car-S1 ET to Chl a. The third lifetime, which extended from ~200 ps to the nanosecond timescale, was attributed to Chl a fluorescence. The 𝜆𝑒𝑥 of 489 nm directly excites the Chl Soret region, whilst excitation at 506 nm shows a pump intensity-dependence. Laser pulse photon density values were ~1014 photons·cm-2·pulse-1 for these datasets. Singlet-singlet annihilation calculations performed on the samples excited at 506 nm provided low annihilation probabilities of 9.0% and 11.5% for a low and high pump intensity, respectively, limiting the possibility of sample photobleaching. Optimization and redevelopment of the experimental setup significantly improved both the data quality and various recorded parameters, concluding that pump-probe spectroscopy was successful on the prepared LHCII trimers. Results acquired and calculations performed correlated with literature, where minimal changes were noticed in the timescales and ET pathways. The robustness of plant systems was confirmed through both excitation-wavelength and intensity dependence. This work paves the way for advanced studies on the role Cars play in non-photochemical quenching (NPQ), a self-protection mechanism of plants against over-illumination; and for the tailoring of artificial light-harvesting antennas based on research conducted on their natural counterparts. / Globale energievereistes het oor die afgelope paar dekades toegeneem, wat die ontwikkeling van alternatiewe energiebronne noodsaaklik maak. Sontegnologieë, geïnspireer deur die primêre sonenergiebergingsproses op aarde, fotosintese, het daarom gewild geword. Die natuurlike fotosintetiese apparaat bestaan uit 'n netwerk van membraangebonde pigment-proteïenkomplekse, met die hoof ligversamelingskompleks in plante (LHCII) wat bestaan uit chlorofil- (Chl) en karotenoïed- (Car) pigmente. Die energie wat deur die pigmente geabsorbeer word, word tussen elektroniese opgewekte toestande op verskillende pigmente op ultravinnige tydskale oorgedra. Hierdie energie word na ʼn fotosintetiese reaksiesentrum gekanaliseer, waar 'n ladingskeiding geïnduseer word en 'n Biobattery sodoende geskep word. Die energie wat in dié battery gestoor is, word gebruik om energieryke chemiese verbindings te vervaardig — wat as brandstof vir die plant dien om sy lewensfunksies te verrig. Tydopgeloste-absorpsie-pomp-tasting-spektroskopie is 'n nuttige tegniek om die dinamika tussen opgewekte toestande te volg. ‘n Voorbeeld van sulke dinamika is die elektroniese opwekking en energie-oordrag tussen die Car- en Chl-pigmente van geïsoleerde LHCII-trimere in spinasieblare. Hierdie metode is gebruik om monsters onder vier verskillende toestande te ondersoek by pompgolflengtes (𝜆𝑒𝑥) van 489 nm (waar hoofsaaklik die Cars Luteïne1 en Neoksantine opgewek word) en 506 nm (vir Cars Luteïne2 en Violaksantine), en pompenergieë van ‘n relatief hoë 800 nJ/puls, of 500 nJ/puls vir elke golflengte. / Dissertation (MSc)--University of Pretoria, 2017. / National Research Foundation (NRF) / Physics / MSc / Unrestricted