The time resolved fluorescence and anisotropy of subtilisins BPN' and Carlsberg.

Time-resolved emission and anisotropy have been measured for the tryptophan (Trp) residues of two closely related subtilisin proteins. The single Trp of subtilisin Carlsberg shows complex lifetime properties, and anisotropy consistent with a fast (ca. 200 ps) segmental motion, on the "wobbling in a cone model" the semi angle is in the range 38 to 47 degrees. The lifetime and anisotropy properties for this single Trp residue suggest that the predominant state is that of an effectively non-emitting statically quenched fluorophore. This fast component is also resolved in the anisotropy of subtilisin BPN' but with relatively low amplitude, due to the dominant emission of the other Trp residues. The diversity of the photophysical properties is not readily correlated with the structure of the proteins, though the observed complexity is consistent with the likely heterogeneity of environment due to the surface location of all the Trp residues.