The modulation of protein kinase C activity by membrane lipid bilayer structure.
The hypothesis that protein kinase C (PKC) activity is sensitive to phospholipid head group interactions was tested using lipid bilayers of defined composition with PKC purified from rat brain. The head group interactions were modulated by varying phosphatidylcholine cis-unsaturation, vesicle curvature, and by the addition of phosphatidylethanolamine and cholesterol. With unilamellar vesicles (including 20 mol% brain phosphatidylserine), increased phosphatidylcholine unsaturation potentiated basal and phorbol ester stimulated PKC activity. By contrast, in the presence of phosphatidylethanolamine, the activity decreased with increasing phosphatidylcholine unsaturation. Weakening phospholipid head group interactions spaces the head group region and increases interstitial water, and this effect was assessed from its effect on the fluorescence intensity of the phospholipid-labeled fluorophore 1-palmitoyl-2-N-(4-nitrobenzo-2-oxa-1,3-diazole)aminohexanoylphosphat idylcholin e (C6-NBD-PC). When the PKC activities with vesicles of varying phosphatidylcholine unsaturation, with and without phosphatidylethanolamine, were plotted as a function of the fluorescence intensity of C6-NBD-PC-labeled vesicles, a biphasic profile was obtained, which had an optimum value of intensity, relating to head group spacing, that corresponded to a maximal enzyme activity. A similar biphasic curve was also found when PKC activities were plotted as a function of published bilayer intrinsic curvature x-ray diffraction data, a parameter closely related to head group spacing. By contrast, no simple relationship was evident between PKC activity and 1,6-diphenyl-1,3,5-hexatriene anisotropy, taken as a measure of lipid order or fluidity. Therefore, increasing the level of phosphatidylcholine unsaturation, phosphatidylethanolamine, or cholesterol either potentiates or attenuates PKC activity, dependent on whether the initial condition is above or below its optimum.
Slater
S J
SJ
Department of Pathology and Cell Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107.
Kelly
M B
MB
Taddeo
F J
FJ
Ho
C
C
Rubin
E
E
Stubbs
C D
CD
D000818:Animals; D001921:Brain / Q000201:enzymology; D005456:Fluorescent Dyes; D007700:Kinetics; D008051:Lipid Bilayers; D008560:Membrane Fluidity; D008823:Micelles; D004676:Myelin Basic Protein / Q000378:metabolism; D010713:Phosphatidylcholines / Q000494:pharmacology; D010714:Phosphatidylethanolamines / Q000494:pharmacology; D010718:Phosphatidylserines / Q000494:pharmacology; D010743:Phospholipids / Q000494:pharmacology*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship; D014961:X-Ray Diffraction
D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D008823:Micelles; D004676:Myelin Basic Protein; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010718:Phosphatidylserines; D010743:Phospholipids; C057561:1-palmitoyl-2-oleoylphosphatidylethanolamine; C044745:1-palmitoyl-2-(6-(N-(7-nitrobenz)-2-oxa-1,3-diazol-4-yl)aminocaproyl)phosphatidylcholine; D011493:Protein Kinase C; C028694:1-palmitoyl-2-oleoylphosphatidylcholine