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 resonance energy transfer from excited tryptophans to pyrene with time-correlated single photon counting. The average lifetime of the excited state of the tryptophans in PKC showed a clear dependence on the mole percentage pDG in micelles in contrast with pyrene-labeled phosphatidylcholine (pPC). The binding data has been analyzed to a simple model which encompasses the size of the micelles and the binding constant of the pDG-PKC complex. From our data, though, these quantities cannot be determined independently. If we have no size information on the micelles we can determine a lower boundary of this quantity compatible with the data. When the micellar size is known, a binding constant for the DG-PKC complex can be extracted. The presented analytical approach can be applied to other systems in which lipid-protein interactions must be quantified.
Bastiaens
P I
PI
Department of Biochemistry, Agricultural University, Wageningen, The Netherlands.
Pap
E H
EH
Borst
J W
JW
van Hoek
A
A
Kulinski
T
T
Rigler
R
R
Visser
A J
AJ
D000818:Animals; D001921:Brain / Q000201:enzymology; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D004075:Diglycerides / Q000737:chemistry / Q000378:metabolism*; D005454:Fluorescence Polarization; D007700:Kinetics; D008823:Micelles; D011493:Protein Kinase C / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D011721:Pyrenes / Q000737:chemistry / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar
D004075:Diglycerides; D008823:Micelles; D011721:Pyrenes; C030984:pyrene; D011493:Protein Kinase C