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Stable Transfection of the Singlet Oxygen Photosensitizing Protein SOPP3: Examining Aspects of Intracellular Behavior

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Protein-encased chromophores that photosensitize the production of reactive oxygen species, ROS, have been the center of recent activity in studies of oxidative stress. One potential attribute of such systems is that the local environment surrounding the chromophore, and that determines the chromophore's photophysics, ideally remains constant and independent of the global environment into which the system is placed. Therefore, a protein-encased sensitizer localized in the mitochondria would arguably have the same photophysics as that protein-encased sensitizer at the plasma membrane, for example. One thus obtains a useful tool to study processes modulated by spatially localized ROS. One ROS of interest is singlet oxygen, O-2(a(1)Delta(g)). We recently developed a singlet oxygen photosensitizing protein, SOPP, in which flavin mononucleotide, FMN, is encased in a re-engineered light-oxygen-voltage protein. One goal was to ascertain how a version of this system, SOPP3, which selectively makes O-2(a(1)Delta(g)), in vitro, behaves in a cell. We now demonstrate that SOPP3 undergoes exacerbated irradiation-mediated bleaching when expressed at either the plasma membrane or mitochondria in stable cell lines. We find that the environment around the SOPP3 system affects the bleaching rate, which argues against one of the key suppositions in support of a protein-encased chromophore.

OriginalsprogEngelsk
TidsskriftPhotochemistry and Photobiology
Vol/bind97
Nummer6
Sider (fra-til)1417-1430
Antal sider14
ISSN0031-8655
DOI
StatusUdgivet - 2021

Bibliografisk note

†This article is part of a Special Issue celebrating the career of Dr. Edward Clennan.

Funding Information:
A grant from the Danish Research Council supported the work described herein. The authors thank (a) Anette Kjems (Aarhus University) for assistance with cloning, protein expression and purification, and (b) Karina Hjorth, Mikkel Roland Holt, and Christian Damgaard (Aarhus University) for assistance with the stable transfections.

Publisher Copyright:
© 2021 American Society for Photobiology

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