OXYGEN PENETRATION AND DIFFUSION INTO MYOGLOBIN REVEALED BY QUENCHING OF ZINCPROTOPORPHYRIN-IX FLUORESCENCE

Oxygen quenching experiments were carried out on zincprotoporphyrin IX reconstituted myoglobin (Mb(Fe-->Zn)) at different temperatures and two solvent viscosities. The data were fit to a dynamic model for quenching of fluorophores in protein interiors previously presented (Biophysical J., 45 (1984) 789-794). The parameters associated with the oxygen entry rate (k(+)), exit rate (k(-)), and migration rate( chi) in the protein were obtained at six temperatures and two viscosities (1 and 8 cp), along with the activation enthalpies associated with the above rates(k(+) and k(-)). The partition coefficient (alpha) was calculated at each temperature along with the free energy, Delta G(0), associated with this partition. The rate parameters (k(+), k(-), chi) and the partition coefficient (alpha) have also been determined for the sample in 40% sucrose (8 cp), to evaluate the effect of bulk solvent viscosities on these values. The steady-state Stern-Volmer quenching plot was calculated using the rate parameters obtained from the analysis (of the dynamic model). Comparison of the Stern-Volmer points obtained using the dynamic model and those obtained experimentally showed excellent agreement.