Figure 2.

Mitofusin 2 disease mutants have retained GTP-binding activity. Wild-type (WT) and disease mutant MFN2 constructs with GFP fused to the N terminus were transfected into HEK 293T cells, and cell lysates were prepared 24 h later. The lysates were incubated with GTP-agarose beads, and bound GFP-MFN2 protein was measured using a fluorimeter. A, GFP–MFN2 binding to GTP-agarose beads was specific, because there is no background binding of GFP alone, and excess GTP (10 mm) was able to competitively block GFP–MFN2 binding to GTP-agarose beads. B, GTP-agarose binding as above using GFP-MFN2 mutants performed in duplicate demonstrates that all of the MFN2 disease mutants bind GTP to a similar extent.