Fig. 3.

CAPS stimulates SNARE complex formation. (A) CAPS stimulates formation of a ≈165 kDa SNARE complex. Incubations were assembled with v- or t-SNARE(5% PIP₂) liposomes or with both in the absence or presence of 1 μM CAPS and incubated for 30 min at 32°. One incubation (asterisk) was boiled before SDS/PAGE. Complexes of ≈165 kDa (arrowhead) were detected with syntaxin antibody. (B) Incubations were conducted with VAMP2- and t-SNARE(5% PIP₂) liposomes for 0 time (lanes 1 and 4) or for 30 min at 32 °C (lanes 2 and 3) or 4 °C (lanes 5 and 6). One micrometer of CAPS was present as indicated (lanes 3 and 6). Complexes of ≈165 kDa (arrowhead) were detected with syntaxin antibody. (C) Incubations with CAPS as in B were conducted for indicated times at 32 °C were analyzed for SDS-resistant SNARE complexes with syntaxin antibody. Intensities of the ≈165 kDa band (arrowhead) were quantified and expressed as fold increase. A representative blot is shown for the 0- to 10-min incubations. One 10-min sample (10*) was boiled before SDS/PAGE. (D) Liposome incubations as in B were conducted for 0 time (lane 1) or 30 min (lanes 2–5) with 1 μM CAPS and 0.1 mM LPC as indicated. SNARE complexes of ≈165 kDa (arrowhead) were detected with syntaxin antibody.