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Hence only relatively slow minute time scale double-kinetic studies could be performed. We have improved on the data density obtained in these pioneering experiments in the following manner: the time required to collect each time slice has been reduced from - 6 0 sec to 1 msec per data set (60,000-fold improvement); the number of photons obtained in the peak of each time-sliced data set was enhanced from -300-400 to 20,000-30,000 counts (~100-fold improvement); the timing resolution was decreased from 102 psec/channel to 1 to 6 psec/channel (-100-fold improvement); Glan-Thompson polarizers were inserted into the detection channel(s) such that double-kinetic anisotropy decays could be measured; and simultaneous multiple-emission wavelength detection was added (up to 12 time-resolved channels).

Ajtai, P. J. K. Ilich, A. Ringler, S. S. Sedarous, D. J. Tort, and T. P. Burghardt, Biochemistry 31, 12431 (1992). 61 B. D. Hamman, A. V. Oleinikov, G. G. Jokhadze, D. E. Bochkariov, R. R. Traut, and D. M. Jameson, J. Biol. Chem. 271, 7568 (1996). [2] DESIGN OF PROFLUORESCENT PROTEASE SUBSTRATES 21 +U Excited State - - i-u G r o u n d State Monomer Dimer F]G. 2. Exciton theory. phores is perpendicular to their transition dipoles. Although previous depictions of rhodamine dimers have indicated face-to-face s t r u c t u r e s , 62-64 m o r e recent data point toward an edge-to-edge alignment between the two dyes.

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