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. 2015 Oct 29;290(50):30053–30065. doi: 10.1074/jbc.M115.665604

FIGURE 7.

FIGURE 7.

The Did2-MIM1 element cannot functionally replace the Ist1 MIM element. A, UV traces for SEC analyses of Ist1(Did2-MIM1) (cyan), Ist1(L168A,Y172A) (green), and Ist1(L168A,Y172A, Did2-MIM1) (green, dashed). The purity of recombinant Ist1 proteins as assessed by SDS-PAGE analysis and Coomassie staining is shown in the right panel. Lane 1, Ist1(Did2-MIM1); lane 2, Ist1(L168A,Y172A,Did2-MIM1). A.U., absorption units. B, limited proteolysis of Ist1 with trypsin. Aliquots were removed at various time points for SDS-PAGE analysis and Coomassie staining. Full-length (FL) Ist1 and the A fragments are indicated. C, ATPase activities of 500 nm Vps4 in the presence of 8 μm Ist1 Did2-MIM1 chimeras with or without L168A,Y172A. Results are presented as mean + S.D. of triplicate experiments, with statistical differences from 500 nm Vps4 alone indicated (***, p < 0.001).