Switch from inhibition to activation of the mitochondrial permeability transition during hematoporphyrin-mediated photooxidative stress.: Unmasking pore-regulating external thiols |
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Authors: | Valeria Petronilli Alessandra Zulian Giulio Jori Giuseppe Tognon Paolo Bernardi |
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Institution: | a C.N.R. Institute of Neurosciences at the Department of Biomedical Sciences, University of Padova, Italy b C.N.R. Institute of Biomedical Technologies at the Department of Biology, University of Padova, Italy c Department of Pharmacology and Anesthesiology/Pharmacology Division, University of Padova, Italy d Bulgarian Academy of Sciences, Institute of Organic Chemistry, Sofia, Bulgaria |
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Abstract: | We have studied the mitochondrial permeability transition pore (PTP) under oxidizing conditions with mitochondria-bound hematoporphyrin, which generates reactive oxygen species (mainly singlet oxygen, 1O2) upon UV/visible light-irradiation and promotes the photooxidative modification of vicinal targets. We have characterized the PTP-modulating properties of two major critical sites endowed with different degrees of photosensitivity: (i) the most photovulnerable site comprises critical histidines, whose photomodification by vicinal hematoporphyrin causes a drop in reactivity of matrix-exposed (internal), PTP-regulating cysteines thus stabilizing the pore in a closed conformation; (ii) the most photoresistant site coincides with the binding domains of (external) cysteines sensitive to membrane-impermeant reagents, which are easily unmasked when oxidation of internal cysteines is prevented. Photooxidation of external cysteines promoted by vicinal hematoporphyrin reactivates the PTP after the block caused by histidine photodegradation. Thus, hematoporphyrin-mediated photooxidative stress can either inhibit or activate the mitochondrial permeability transition depending on the site of hematoporphyrin localization and on the nature of the substrate; and selective photomodification of different hematoporphyrin-containing pore domains can be achieved by fine regulation of the sensitizer/light doses. These findings shed new light on PTP modulation by oxidative stress. |
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Keywords: | ANT adenine nucleotide translocase CsA cyclosporin A Cyp-D cyclophilin D Cu(OP)2 copper-o-phenanthroline Cys cysteine DTT dithiothreitol EGTA [ethylenebis(oxoethylenenitrilo)] tetraacetic acid His histidine HP hematoporphyrin IX IMM inner mitochondrial membrane MBM+ trimethylammonium monobromobimane MOPS 4-morpholinepropanesulfonic acid 1O2 singlet oxygen OMM outer mitochondrial membrane PhAsO phenylarsine oxide PT permeability transition PTP permeability transition pore RCR respiratory control ratio ROS reactive oxygen species TEM transmission electron microscopy VDAC voltage-dependent anion channel |
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