FIGURE 5.

Binding of α-synuclein and transglutaminases to LV. 10 pmol of α-synuclein was added to varying amounts of LV from 15% phosphatidylserine, 55% phosphatidylcholine, and 30% cholesterol at room temperature (solid circles) and 37 °C (open circles). The not membrane-bound fraction of protein was measured from centrifugal ultrafiltrate. A, warming LV/α-synuclein mixtures to 37 °C (triangles) results in increased protein binding, which is preserved at the lower temperature thereafter. This phenomenon indicates that membrane fluidity is needed to fit interacting PS and α-synuclein residues together. B, calcium ions increase α-synuclein association to PS-containing LV. The ratio of synuclein/PS (mol/mol) is plotted; error bars show 95% confidence intervals. C, binding of TG1 to LV is independent of calcium concentration, whereas TG2 and TG5 associate with LV in a calcium ion-dependent manner. In the presence of LV, micromolar calcium ion concentrations elicit detectable TG activation for TG2 and TG5 (D); however, in solution these concentrations do not activate them (inset in D). The patterning of columns showing different free Ca²⁺ concentrations is the same as on C. TG1 is fully active at the lowest tested calcium concentration, whereas TG3 does not associate with LV and neither is its activity affected by LV. TG activity was measured by assaying p-nitrophenyl acetate hydrolysis by photometry. Points indicate means of five independent assays.