X-ray reflectivity measurements are used to determine the configuration of the C2 domain of protein kinase C
α (PKC
α-C2) bound to a lipid monolayer of a 7:3 mixture of 1-stearoyl-2-oleoyl-
sn-glycero-3-phosphocholine and 1-stearoyl-2-oleoyl-
sn-glycero-3-phosphoserine supported on a buffered aqueous solution. The reflectivity is analyzed in terms of the known crystallographic structure of PKC
α-C2 and a slab model representation of the lipid layer. The configuration of lipid-bound PKC
α-C2 is described by two angles that define its orientation,
θ = 35° ± 10° and
φ =210° ± 30°, and a penetration depth (=7.5 ± 2 Å) into the lipid layer. In this structure, the
β-sheets of PKC
α-C2 are nearly perpendicular to the lipid layer and the domain penetrates into the headgroup region of the lipid layer, but not into the tailgroup region. This configuration of PKC
α-C2 determined by our x-ray reflectivity is consistent with many previous findings, particularly mutational studies, and also provides what we believe is new molecular insight into the mechanism of PKC
α enzyme activation. Our analysis method, which allows us to test all possible protein orientations, shows that our data cannot be explained by a protein that is orientated parallel to the membrane, as suggested by earlier work.
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