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Studies On The Structure And Function Of Light Harvesting Complexes Ⅱ(LH2) And Photosynthetic Reaction Centers (RC) Of Rhodobacter Sphaeroides

Posted on:2006-02-10Degree:DoctorType:Dissertation
Country:ChinaCandidate:Y LiuFull Text:PDF
GTID:1100360152499422Subject:Botany
Abstract/Summary:PDF Full Text Request
Light harvesting complexes II (LH2) and Photosynthetic reaction centers (RC) of Rhodobacter sphaeroides were investigated to study the stucture and function by the biochemical and biophysical methods. The results obtained here were summarized as follows: 1. Acquirement and Characterisation of a Carotenoid Mutant (GM309) of Rhodobacter sphaeroides 601 A green mutant was obtained among chemical induced mutants of Rhodobacter sphaeroides 601 (RS601) and named as GM309. A blue shift of 20 nm of the carotenoid absorption spectrum was found in the Light-harvesting complex II (LH2) of GM309. Different from LH2 of RS601, it was found that the carotenoids in GM309-LH2 changed to be neurosporene by mutation. Neurosporene lacks a conjugate double bond, compared with the spheroidene in RS601-LH2 which has ten conjugate double bonds. As shown by absorption and circular dichroism spectroscopy, the overall structure of GM309-LH2 is little affected by this change. From fluorescence emission spectra, it is found that GM309-LH2 can transfer energy from carotenoids to Bchl-B850 without any change in efficiency. But the efficiency of energy transfer from B800 to B850 in GM309-LH2 is decreased to be 42% of that of the native. This work would provide a novel method to investigate the mechanism of excitation energy transfer in LH2. 2.Elimination of polarity in the carotenoid terminus promotes the exposure of B850 binding sites (Tyr 44, 45) and ANS-mediated energy transfer in LH2 complexes of Rhodobacter sphaeroides Carotenoids in GM309-LH2 were identified as elimination of polarity group in one terminus, compared to that in native-LH2. After LH2 complexes were treated with hydrophobic fluorescence probe(ANS), new energy transfer pathways from ANS or tryptophan to carotenoids were discovered in both native-LH2 and GM309-LH2. The carotenoid fluorescence intensity of GM309-LH2 was greater than that of native-LH2, suggesting that the elimination of polarity in carotenoid in one terminusmay increase hydrophobic residues to bind ANS probe. The further finding that two a-tyrosines (a-Tyr 44, 45, B850-binding sites) in each a-apoprotein of GM309-LH2 were more easily modified than those of native-LH2 by N-acetylimidazole (NAI), indicates that the elimination of polarity in the carotenoid terminus increases the exposure of a-Tyr 44, 45 to solution. 3. The observation of excited-state dynamical evolution in light-harvesting complex(LH2) from Rhodobacter sphaeroides 601 With selective excitation around BChl-B800 and BChl-B850 absorption bands, we observed the evolution of excited-state dynamics in LH2 from Rhodobacter sphaeroides 601. The dynamical traces demonstrate a dominant excited-state absorption (ESA) followed concomitantly by an ultrafast transmission increase and decay with pulse-width limited time scale at 818 nm and 828 nm excitation. The ESA occurring prior to excitonic thermalization or ground-state bleach was observed at 840 nm as well. These experimental results indicate the competition between the transition from excitonic states to higher-lying excited states and interexciton relaxation, which are of physical significance for understanding excitation transfer and related mechanisms in LH2. ? 2002 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.4. The Excitation Relaxation in Chemically Modified Photosynthetic Reaction Center from Rb. sphaeroides 601 The ultrafast excitation relaxation in sodium borohydride treated reaction center of Rb. sphaeroides 601 was investigated with selective excitation. From the femtosecond (fs) pump-probe measurement at 790 nm, the excitation relaxation demonstrates a biexponential decay with time constants of about 200 fs and 1.4 ps, respectively. By comparison with the result from sodium ascorbate pretreated modified RS601, it could be concluded that the dynamical trace at 790 nm mainly originates from the contribution of accessory bacteriochlorophyll in active side, and the electrochromic shift arising from the induced positive charge on special pair primarily affects the absorption band in red-region of accessory bacteriochlorophyll in RS601. With direct excitation of special pair, the charge separation and subsequent electron transfer were observed in borohydride modified RS601. The 2.8 ps component was ascribed to the charge separation and electron transfer from P* to HA. From the dynamical traces at 790, 800 and 818 nm, the ultrafast energy relaxation from excited accessory bacteriochlorophyll in active side is consistent with two-step energy transfer mechanism. This dynamical observation in...
Keywords/Search Tags:Light-harvesting complex II, photosynthetic reaction center, green mutant, ultrafast dynamics, energy transfer, chemically modification
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