The proton activity at cryogenic temperatures--a possible influence on the spin state of the heme iron of cytochrome P-450cam in supercooled buffered solutions.
The electronic absorption spectra for camphor-bound cytochrome P-450cam have been analysed in the temperature range between 78 K and 298 K. The well-known high-spin/low-spin equilibrium has been detected between 298 K and 220 K. Depending on the cooling rate, below 220 K a new species was found in the absorption spectra. In contrast, the electronic absorption spectra for camphor-free cytochrome P-450cam between 78 K and 295 K show no significant spectral changes. The conversion between the spin states of camphor-bound cytochrome P-450cam and the appearance of the new species do not correspond to the temperature-induced change in the paH value of the aqueous glycerol mixture containing phosphate or cacodylate buffer (paH 7.0). For this study a spectroscopic procedure for the determination of the temperature dependence of the paH value of the solvent for the range 78-295 K is presented using dyes as pH-indicators. It is shown that the state of the acid-base equilibrium frozen in is strongly dependent on the cooling rate of the mixture.
Schulze
H
H
Max Delbrück Centre for Molecular Medicine, Berlin-Buch, Germany.
Ristau
O
O
Jung
C
C
D002021:Buffers; D019475:Camphor 5-Monooxygenase; D003080:Cold Temperature; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry*; D006418:Heme; D006863:Hydrogen-Ion Concentration; D007501:Iron; D006899:Mixed Function Oxygenases / Q000737:chemistry*; D011487:Protein Conformation; D011522:Protons; D013053:Spectrophotometry
D002021:Buffers; D011522:Protons; D006418:Heme; D003577:Cytochrome P-450 Enzyme System; D007501:Iron; D006899:Mixed Function Oxygenases; D019475:Camphor 5-Monooxygenase