Increases in peroxide formation by the Photosystem II oxygen evolving reactions upon removal of the extrinsic 16, 22 and 33 kDa proteins are reversed by CaCl2 addition |
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| Authors: | Warwick Hillier Tom Wydrzynski |
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| Affiliation: | (1) Research School of Biological Sciences, The Australian National University, 2601 Canberra, ACT, Australia |
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| Abstract: | This communication introduces a new spectrophotometric assay for the detection of peroxide generated by Photosystem II (PS II) under steady state illumination in the presence of an electron acceptor. The assay is based on the formation of an indamine dye in a horseradish peroxidase coupled reaction between 3-(dimethylamino)benzoic acid and 3-methyl-2-benzothiazolinone hydrazone. Using this assay, we found that as the O2 evolution activity of PS II-enriched membrane fragments is decreased by treatments which cause the dissociation of the 33 and/or 23 and 16 kDa extrinsic proteins (i.e., CaCl2-washing, NaCl-washing, lauroylcholine-treatment and ethylene glycol-treatment), light-induced peroxide formation increases. Both the losses of O2 evolution and increases in peroxide formation seen under these conditions are reversed by CaCl2 addition, indicating that the two activities originate from the water-splitting site. However, the increased rates of peroxide formation do not quantitatively match the losses in O2 evolution activity. We suggest that a rapid consumption of the peroxide takes place via a catalase/peroxidase activity at the water-splitting site which competes with both the O2 evolution and peroxide formation reactions. The observed peroxide formation is interpreted as arising from enhanced water accessibility to the catalytic site upon perturbation of the extrinsic proteins which then leads to alternate water oxidation side reactions.Abbreviations Chl chlorophyll - DCBQ 2,6-dichloro-p-benzoquinone - DCMU 3-(3,4-dichloro)-1,1-dimethylurea - DCPIP 1,6-dichlorophenolindophenol - DMAB 3-(dimethylamino)benzoic acid - DMBQ 2,6-dimethyl-p-benzoquinone - DPC diphenylcarbazide - HEPES 4-(2-hydroxyethyl)-1-piperazinesulfonic acid - HMD HRP, MBTH, DMAB - HRP horseradish peroxidase - LCC lauroylcholine chloride - MBTH 3-methyl-2-benzothiazolinone hydrazone - MES 4-morpholinoethanesulfonic acid |
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| Keywords: | calcium effects extrinsic proteins O2 evolution peroxide formation Photosystem II |
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