A working model of the proteins and membranes as assembled prior to vacuole or proteoliposome fusion. From Zick et al. (2015).
(A) The fusion of Ypt7(GTP):R-SNARE proteoliposomes with Ypt7(GTP):QaQb-SNARE proteoliposomes with added Qc-SNARE and HOPS is stimulated by Sec17, Sec18, and ATP or ATPγS. (B) ATP hydrolysis is …
Source data file (Excel) for Figure 2A.
Source data file (Excel) for Figure 2B.
In panel A, the relative fusion after 10 min is also shown for reactions with HOPS, Sec17, Sec18, and ATP or ATPγS.
Fusion conditions were as described in Materials and methods. (A) Fusion with ATPγS. (B) Fusion with ATP. Sec17 was wild type (wt) or had the mutations F21S,M22S (FSMS), K159E,K163E (KEKE), or …
Source data file (Excel) for Figure 3 Parts A, B, and C.
Blue bars are for incubations from part A, with ATPγS, and black bars from part B, with ATP.
(A) The sSNAREs (deprived of their trans-membrane anchor domains) were mixed with Sec17 and protein-free liposomes, incubated, and the liposomes and bound proteins were isolated by floatation, as …
Data are from Figure 3A and B, which were part of the same experiment.
Data are from Figure 3A and B, which were part of the same experiment.
The transmembrane domain from the Qb-SNARE Vti1 was joined to the N-terminus of Sec17 (wt) or Sec17 F21S,M22S (FSMS) to give TM-Sec17 (wt) and TM-Sec17 FSMS. (A) No Sec17, (B) TM-Sec17 (wt), or C). …
Source data file (Excel) for Figure 4 Parts A, B, and C.
Proteoliposomes bore TM-Sec17 wild-type or TM-Sec17 F21S,M22S as indicated, and reactions had HOPS, Sec18, Sec18 with ATP, or Sec18 with ATPγS as indicated.
(A) Fusion is lost without SNARE zippering unless ATP is replaced by ATPγS. The fusion of proteoliposomes with Ypt7:GTP and either the R-SNARE or QaQb-SNAREs in the presence of Sec17 and Sec18 was …
Source data file (Excel) for Figure 5A.
Source data file (Excel) for Figure 5B.
Source data file (Excel) for Figure 5C.
(A) Fusion reactions were performed with ATPγS as in Figure 3A, but with Qc-3Δ instead of Qc. (B) Fusion reactions were performed as in part A, but with ATP instead of ATPγS. Sec17 was wild type …
Source data file (Excel) for Figure 6 Parts A and B.
Source data file (Excel) for Figure 6C.
Fusion incubations were performed with Ypt7:GTP, 1R-SNARE proteoliposomes and Ypt7:GTP, QaQb-SNARE proteoliposomes in the presence of HOPS, Sec17, Sec18, ATPγS, and Qc-3Δ as described in Materials …
Source data file (Excel) for Figure 7.
Open bars are the average rate for the first 20 min, filled bars are the average rates for the first 6 min after the restoration of the missing component(s) at 20 min. One outlier data point was …