Time-resolved fluorescence of proteins and model systems

TIME-RESOLVED FLUORESCENCE OF PROTEINS AND MODEL SYSTEMSJ.B. Alexander Ross and Maurice R. EftinkDepartment of Biochemistry, Mount Sinai School of Medicine, New York, New York 10029 andDepartment of Chemistry, University of Mississippi, University, Mississippi 38677HISTORICAL BACKGROUNDThe history of protein luminescence is more than a century old. Konev,1 in a review of resultsfrom the nineteenth century, cites the observations by Stokes of the fluorescence of horns and nails,by Regnault and Sechenov of the fluorescence of the crystalline lens and skin, and by Soret of theviolet fluorescence of myosin. Numerous other examples of this phenomenon were described bymany different investigators during the period between the late 1920s to the early 1950s. In general,there is no clear distinction in most of these reports whether the observed luminescence was due tophosphorescence or to fluorescence. It appears, on one hand, that because of the time delay betweenexcitation and detectection, some workers were detecting phosphorescence. On the other hand, itis possible that in fact others were observing the low-energy tail of tryptophan fluorescence, whichextends into the range of visible wavelengths above 400 nm.The review on fluorescence polarization of solutions by Gregorio Weber,2 published in 1953,marks a critically important time in the history of protein fluorescence. In this review, Weber clearlyset the direction for future thinking about the nature of protein fluorescence. He pointed out thatthe ultraviolet absorption spectra of proteins at wavelengths greater than 250 nm was due largely to