Figure 2.

Conditions that alter the abundance of SNARE complexes also alter the Sec1p-Ssop interaction. (A) The interaction of Ssop with either MYC-Sec1p or Sncp is altered in sec mutants. SEC+ (NY1689), sec4-8 (NY1690), and sec18-1 (NY1691) MYC-SEC1 strains were shifted to 37°C for 10 min. The Ssop antibody was used to detect Ssop from the anti-Sncp and the anti-MYC IPs. (B) Sec18p disassembles SNARE complexes and disrupts the Sec1p-Ssop interaction. Lysates of SEC+ (NY1689) and sec18-1 (NY1691) MYC-SEC1 strains were prepared either with ATP (+) or without ATP (−), and the anti-MYC or anti-Sncp IPs were probed for coprecipitated Ssop. When ATP was required, NaN₃ and NaF were omitted from the wash buffer, and EDTA was replaced by an ATP-regeneration system (10 μg/ml creatine kinase, 5 mM creatine phosphate, 1 mM ATP, and 1 mM MgCl₂) in the IP buffer.