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Biochemical and spectroscopic study of the structure and function of higher plant PSI

Berry, Matthew Carl; (1995) Biochemical and spectroscopic study of the structure and function of higher plant PSI. Doctoral thesis (Ph.D.), University College London (United Kingdom). Green open access

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Abstract

This thesis reports structural and functional studies of the photosystem I reaction centre of higher plants. The main techniques used were reaction centre preparation by detergent fractionation and modification by chaotrope treatment, optical and pulsed electron spin polarised kinetic spectroscopies at room and at cryogenic temperatures; continuous wave and time - resolved (ns time scale) pulsed electron spin resonance spectroscopies, and electron spin envelope echo modulation spectroscopy. The kinetic behaviour of spinach PSI particles was examined where the iron - sulphur clusters were chemically prereduced at room temperature and at 4 K. A flash - induced electron spin polarised signal was observed in the out - of - phase detection channel using a short microwave pulse sequence, which decayed with a rate constant of 23 [mu]s. Using microwave pulses of longer duration a spin polarised signal was observed in the in - phase channel decaying with the same rate constant as the former and spectrally identical to the spin polarised signal attributed to the P700+A1-- radical pair. It is concluded that at 4 K forward electron transfer from A1, is inhibited and that the observed spin polarised signal decays by a charge recombination and that the phasing of this signal depends on the characteristics of the pulse sequence used in detection. At room temperature the flash - induced out - of - phase spin polarised signal was found to decay with a rate constant of 130 ns +/_ 50 ns and its decay was accompanied by the rise of a second spin polarised signal attributed to the radical pair, P700+FeS-- which in turn decays with a rate constant of 2 [mu]s. When the iron - sulphur clusters, Fe-SAB, were extracted the room temperature kinetic behaviour of the PSI core particles was largely unaltered relative to intact particles. When the iron - sulphur cluster, Fe-Sx, was extracted as well, the decay of the out - of - phase signal was slowed down to give a rate constant of 1.3 [mu]s and the in - phase signal was abolished. This demonstrates that in intact PSI particles at room temperature electron transfer takes place from A1to the next acceptor and establishes unambiguously that this acceptor is the iron - sulphur cluster, Fe-Sx and means that the "200 ns" kinetic widely reported in the literature can be attributed to the reaction A1—FeSx—[greater-than]A1FeSx–.The same measurements were also carried out on PSI reaction centres prepared from the cyanobacterium, Synechocystis, and very similar kinetic behaviour was observed, implying that there is structural and mechanistic conservation between the two species. The spatial arrangement of the redox components of spinach PSI relative to the iron - sulphur cluster, FeSx, was studied using pulsed saturation recovery experiments. The strength of the magnetic interaction between the reduced non-metal radicals and the iron sulphur cluster, Fe-Sx, quantified by measuring the intrinsic and enhanced spin lattice relaxation rates for each radical species. From these measurements we propose a model of PSI which resembles the 6 [angstrom] crystal structure and the photovoltage model, i.e. a branched arrangement of acceptors. Using a mathematical model we estimated the distances separating the electron acceptors of PSI.

Type: Thesis (Doctoral)
Qualification: Ph.D.
Title: Biochemical and spectroscopic study of the structure and function of higher plant PSI
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest.
Keywords: (UMI)AAI10017786; Biological sciences
URI: https://discovery.ucl.ac.uk/id/eprint/10098491
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