Details
| Original language | English |
|---|---|
| Pages (from-to) | 1170-1181 |
| Number of pages | 12 |
| Journal | Free radical research |
| Volume | 52 |
| Issue number | 10 |
| Publication status | Published - Oct 2018 |
| Externally published | Yes |
Abstract
Hydrogen peroxide (H2O2) plays an important role in various biological processes in numerous organisms. Depending on the concentration and the distribution within the cell, it can act as stressor or redox signalling molecule. To analyse the effects of H2O2 and its diffusion within the cell we developed the new genetically encoded photosensitizer KillerRed-SOD1 which enables a light-induced spatially and temporally controlled generation of H2O2 in living cells. The KillerRed-SOD1 is a fusion protein of the photosensitizer KillerRed (KR) and the cytosolic superoxide dismutase isoform 1 (SOD1) connected by a helix-forming peptide linker. Light irradiation at a wavelength of 560 nm induced superoxide radical formation at the KR domain which was transformed to H2O2 at the SOD1 domain. H2O2 was specifically detected under live cell conditions using the fluorescent sensor protein HyPer. Genetically encoded photosensitizers have the advantage that appropriate tag sequences can determine the localisation of the protein within the cell. Herein, it was exemplarily shown that the peroxisomal targeting sequence 1 directed the photosensitizer KR-SOD1 to the peroxisomes and enabled H2O2 formation specifically in these organelles. In summary, with the photosensitizer KR-SOD1 a new valuable tool was established which allows a controlled intracellular H2O2 generation for the analysis of H2O2 effects on a subcellular level.
Keywords
- Animals, Cell Death, Genetic Engineering, HEK293 Cells, Humans, Hydrogen Peroxide/chemistry, Light, Photosensitizing Agents/chemistry, Rats, Superoxide Dismutase-1/genetics, HyPer, hydrogen peroxide, Genetically encoded photosensitizer, superoxide dismutase, KillerRed, oxidative stress, β cell line
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Biochemistry
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In: Free radical research, Vol. 52, No. 10, 10.2018, p. 1170-1181.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Light-induced intracellular hydrogen peroxide generation through genetically encoded photosensitizer KillerRed-SOD1
AU - Laporte, Anna
AU - Nordenbrock, Anke
AU - Lenzen, Sigurd
AU - Elsner, Matthias
N1 - Funding information: The excellent technical assistance of Martin Wirth, Britta Less, and Anke Possler is gratefully acknowledged. We would like to acknowledge the assistance of the Cell Sorting Core Facility at the Hannover Medical School supported in part by Braukmann-Wittenberg-Herz-Stiftung and Deutsche Forschungsgemeinschaft. The work was supported by Deutsche Forschungsgemeinschaft, GRK 1947/1 and the Deutsche Diabetes Gesellschaft.
PY - 2018/10
Y1 - 2018/10
N2 - Hydrogen peroxide (H2O2) plays an important role in various biological processes in numerous organisms. Depending on the concentration and the distribution within the cell, it can act as stressor or redox signalling molecule. To analyse the effects of H2O2 and its diffusion within the cell we developed the new genetically encoded photosensitizer KillerRed-SOD1 which enables a light-induced spatially and temporally controlled generation of H2O2 in living cells. The KillerRed-SOD1 is a fusion protein of the photosensitizer KillerRed (KR) and the cytosolic superoxide dismutase isoform 1 (SOD1) connected by a helix-forming peptide linker. Light irradiation at a wavelength of 560 nm induced superoxide radical formation at the KR domain which was transformed to H2O2 at the SOD1 domain. H2O2 was specifically detected under live cell conditions using the fluorescent sensor protein HyPer. Genetically encoded photosensitizers have the advantage that appropriate tag sequences can determine the localisation of the protein within the cell. Herein, it was exemplarily shown that the peroxisomal targeting sequence 1 directed the photosensitizer KR-SOD1 to the peroxisomes and enabled H2O2 formation specifically in these organelles. In summary, with the photosensitizer KR-SOD1 a new valuable tool was established which allows a controlled intracellular H2O2 generation for the analysis of H2O2 effects on a subcellular level.
AB - Hydrogen peroxide (H2O2) plays an important role in various biological processes in numerous organisms. Depending on the concentration and the distribution within the cell, it can act as stressor or redox signalling molecule. To analyse the effects of H2O2 and its diffusion within the cell we developed the new genetically encoded photosensitizer KillerRed-SOD1 which enables a light-induced spatially and temporally controlled generation of H2O2 in living cells. The KillerRed-SOD1 is a fusion protein of the photosensitizer KillerRed (KR) and the cytosolic superoxide dismutase isoform 1 (SOD1) connected by a helix-forming peptide linker. Light irradiation at a wavelength of 560 nm induced superoxide radical formation at the KR domain which was transformed to H2O2 at the SOD1 domain. H2O2 was specifically detected under live cell conditions using the fluorescent sensor protein HyPer. Genetically encoded photosensitizers have the advantage that appropriate tag sequences can determine the localisation of the protein within the cell. Herein, it was exemplarily shown that the peroxisomal targeting sequence 1 directed the photosensitizer KR-SOD1 to the peroxisomes and enabled H2O2 formation specifically in these organelles. In summary, with the photosensitizer KR-SOD1 a new valuable tool was established which allows a controlled intracellular H2O2 generation for the analysis of H2O2 effects on a subcellular level.
KW - Animals
KW - Cell Death
KW - Genetic Engineering
KW - HEK293 Cells
KW - Humans
KW - Hydrogen Peroxide/chemistry
KW - Light
KW - Photosensitizing Agents/chemistry
KW - Rats
KW - Superoxide Dismutase-1/genetics
KW - HyPer
KW - hydrogen peroxide
KW - Genetically encoded photosensitizer
KW - superoxide dismutase
KW - KillerRed
KW - oxidative stress
KW - β cell line
UR - http://www.scopus.com/inward/record.url?scp=85059190730&partnerID=8YFLogxK
U2 - 10.1080/10715762.2018.1540042
DO - 10.1080/10715762.2018.1540042
M3 - Article
C2 - 30350732
VL - 52
SP - 1170
EP - 1181
JO - Free radical research
JF - Free radical research
SN - 1071-5762
IS - 10
ER -