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Influence of the Physical State of the Membrane on the Enzymatic Activity and Energy of Activation of Protein Kinase C α

1999; American Chemical Society; Volume: 38; Issue: 24 Linguagem: Inglês

10.1021/bi983062z

1943-295X

Antonia M. Jiménez‐Monreal, Francisco J. Aranda, Vicente Micol, Pilar Sánchez‐Piñera, Ana de Godos, Juan C. Gómez‐Fernández,

Surfactants and Colloidal Systems

The activation of protein kinase C α was studied by using a lipid system consisting of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoserine (POPS) (molar ratio 4:1) and different proportions of 1-palmitoyl-2-oleoyl-sn-glycerol (POG). The phase behavior of the lipidic system was characterized by using differential scanning calorimetry and 31P NMR, and a phase diagram was elaborated. The results suggested the formation of two diacylglycerol/phospholipid complexes, one at 15 mol % of POG and the second at 30 mol % of POG. These two complexes would define the three regions of the phase diagram: in the first region (concentrations of POG lower than 15 mol %) there is gel−gel immiscibility at temperatures below that of the phase transition between C1 and pure phospholipid, and a fluid lamellar phase above of the phase transition. In the second region (between 15 and 30 mol % of POG), gel−gel immiscibility between C1 and C2 with fluid−fluid immiscibility was observed, while inverted hexagonal HII and isotropic phases were detected by 31P NMR. In the third region (concentrations of POG higher than 30 mol %), gel−gel immiscibility seemed to occur between C2 and pure POG along with fluid−fluid immiscibility, while an isotropic phase was detected by 31P NMR. When PKC α activity was measured, as a function of POG concentration, maximum activity was found at POG concentrations as low as 5−10 mol %; the activity slighty decreased as POG concentration was increased to 45 mol % at 32 °C (above Tc) whereas activity did not change with increasing concentrations of POG at 5 °C (below Tc). When the activity was studied as a function of temperature, at different POG concentrations, and depicted as Arrhenius plots, it was found that the activity increased with increasing temperatures, showing a discontinuity at a temperature very close to the phase transition of the system and a lower activation energy at the upper slope of the graph, indicating that the physical state of the membrane affected the interaction of PKC α with the membrane.