PORPHYRIN PHOTOSENSITIZATION OF PROTEINS IN CELL MEMBRANES AS STUDIED BY SPIN-LABELLING AND BY QUANTIFICATION OF DTNB-REACTIVE SH-GROUPS |
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Authors: | Johan Moan Arnt I. Vistnes |
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Affiliation: | Norsk Hydros Institute for Cancer Research, The Norwegian Radium Hospital, Montebello 0310, Oslo 3, Norway;Institute of Physics, University of Oslo, Blindern, Oslo 3, Norway |
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Abstract: | Abstract— Human cells of the line NHIK 3025 were exposed to hematoporphyrin derivative (Hpd) and light and analysed with respect to; (i) the mobility of membrane proteins as determined by electron spin resonance measurements of a protein-bound spin label, (ii) fluorescence excitation spectra, (iii) relative number of DTNB-reactive SH-groups on their surface and in sonicated cell homogenates, (iv) survival, and (v) morphologic appearance as seen by ordinary phase contrast microscopy. A significant fraction of the porphyrins bound to the outer cell membrane was in close contact with proteins. 5,5'-Dithiobis-2-nitrobenzoic acid reactive SH-groups on the outer cell membrane were very sensitive to the treatment with Hpd + light and were degraded according to non-exponential kinetics. When the cells were irradiated after spin labelling, the labelled proteins became less mobile during the irradiation, indicating protein cross linking. Irradiation before spinlabelling resulted in a selective degradation of low-mobility proteins. |
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