Application Notes

Interactions of liposome embedded SNARE proteins

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Material and Methods Assay conditions SNARE proteins were expressed and purified as described (Schuette et al. 2004). Proteoliposomes used in this experiment were prepared as described in 20 mM HEPES, pH 7.4, with 150 mM KCl (Pobbati et al. 2006). Lipid composition was in 5:2:2:1 ratio of brain L-α-phosphatidylcholine, L-α- phosphatidylethanolamine (PE), L-α- phosphatidylserine and cholesterol (Avanti). Protein:lipid ratio was 1:4000. For the experiment described here, a 20 mM HEPES, 150 mM KCl at pH 7.4 buffer has been used. The measurement was performed in standard treated capillaries (Cat#K002). Instrumentation The measurements were done on NanoTemper Monolith NT.015 and NT.115 instruments. The measurement was performed at 40 % LED and 40 % MST power, Laser-On time was 30 sec, Laser-Off time 5 sec. References Schütte et al., Determinants of liposome fusion mediated by synaptic SNARE proteins. Proc Natl Acad Sci USA 2004; 101;2858–2863. Pobbati et al., N- to C-terminal SNARE complex assembly promotes rapid membrane fusion Science 2006 313;673–676 © 2011 NanoTemper Technologies GmbH

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