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Low-temperature phototransformation in mesoporphyrin-IX-substituted horseradish peroxidase

Low-temperature phototransformation in mesoporphyrin-IX-substituted horseradish peroxidase. Artur Suisalu a) and Judit Fidy b) a) Institute of Physics, University of Tartu, Riia 142, 51014 Tartu, Estonia

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Low-temperature phototransformation in mesoporphyrin-IX-substituted horseradish peroxidase

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  1. Low-temperature phototransformationin mesoporphyrin-IX-substituted horseradish peroxidase Artur Suisalua) and Judit Fidyb) a)Institute of Physics, University of Tartu, Riia 142,51014 Tartu, Estonia b)Institute of Biophysics and Radiation Biology, Semmelweis University, Budapest H-1444, Hungary

  2. Contents: • Photobleaching at nonselective excitation • ODMR in photoproduct state • Selectively excited phototransformation • Temperature cycling • Pressure effects Acknowledgment: This research was supported by the Estonian and Hungarian Academies of Sciences. Personally, we thank: Prof. Jaak Kikas, Dr. Herenyi Levente, Dr. Koit Mauring Dr. Arlentin Laisaar and Dr. Anatoli Kuznetsov A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

  3. Horseradish peroxidase: a medium-size protein (308 residues) mesoporphyrin-IX A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

  4. 0,0-bands of mesoporphyrin-IX in apoHRP at 5K excitation spectrum after cooling from RT emission spectrum after irradiation in the Soret band A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

  5. Phototransformation in the 0,0-band of MP-IX-HRP under nonselective excitation in the Soret band Area of the three bands does not conserve. A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

  6. ODMR in photoproduct states B2 and B3 • ODMR spectra are additionally broadened due to “doubling” of MW resonances; • Relaxed triplet state is not involved in NH-tautomerization; • X-Y-asymmetry of MP-IX is smaller in the intermediate state of B2 than in B3. A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

  7. Thermal recovery of the selectively photobleached band B1 A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

  8. Revealing a new state B5 by selective photobleaching A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

  9. Thermal recovery of the selectively photobleached band B5 A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

  10. 2-dimensional excitation-emission spectrum of MP-IX-HRP 1D section of 2D spectrum along the line nex-nem=W=const. gives inhomogeneous distribution function A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

  11. Pressure effect on the inhomogeneous distribution function • Pressure induced repopulation of tautomers occurs at low temperature of the pressuration, • The educt state (B1) is less compressible than product state (B3), A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

  12. Pressure shifts: nP-n0=2(n0- nvac)kP, and broadening: sP=(1+2kP)s A.Suisalu & J. Fidy, Low-temperature phototransformation in MP-IX-HRP

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