Basic Fluorescence Principles I

The polarization observed in buffer alone was 0.151 while the limiting polarization obtained from the intercept on the Y-axis was 0.225, which is the same value one obtains for upon excitation of protoporphyrin IX in glycerol at low temperatures. From the Perrin equation:

and knowing the lifetime of 16.9 ns, one can calculate a rotational relaxation time of 96 ns for the protein-porphyrin complex:

For a spherical protein of 44,000 daltons and assuming a partial specific volume of 0.74 and 0.3 ml/mg for the hydration, one can then calculate:

r = 96 ns

ro = (3)(0.01)(44000)(0.74+0.3)/(8.31x107)(293) = ~56 ns

Thus it appears as if this protein is non-spherical

Polarization VIII