THE INTERACTION OF PYRENE-LABELED DIACYLGLYCEROL WITH PROTEIN-KINASE-C IN MIXED MICELLES

The binding of protein kinase C (PKC) to pyrene-labeled diacylglycerol (pDG) has been studied in a mixed micellar system by monitoring reson ance energy transfer from excited tryptophans to pyrene with time-corr elated single photon counting. The average lifetime of the excited sta te of the tryptophans in PKC showed a clear dependence on the mole per centage pDG in micelles in contrast with pyrene-labeled phosphatidylch oline (pPC). The binding data has been analyzed to a simple model whic h encompasses the size of the micelles and the binding constant of the pDG-PKC complex. From our data, though, these quantities cannot be de termined independently. If we have no size information on the micelles we can determine a lower boundary of this quantity compatible with th e data. When the micellar size is known, a binding constant for the DG -PKC complex can be extracted. The presented analytical approach can b e applied to other systems in which lipid-protein interactions must be quantified.