Figure 2: ATOM14 is an ATOM complex subunit with a cytosol-facing N-terminus. | Scientific Reports

Figure 2: ATOM14 is an ATOM complex subunit with a cytosol-facing N-terminus.

From: Mitochondrial protein import - Functional analysis of the highly diverged Tom22 orthologue of Trypanosoma brucei

Figure 2

(A) Digitonin-extracted crude mitochondrial fractions of procyclic T. brucei cell lines expressing N- or C-terminally c-Myc-tagged ATOM14, respectively, were subjected to co-immunprecipitations using an anti-c-Myc antiserum. Immunoblots containing 5% input (In) and 100% eluate (IP) were probed for the presence of the tagged proteins and the indicated ATOM complex subunits. VDAC serves as a control. (B) C-terminally c-Myc-tagged ATOM14 was immunoprecipitated using crude mitochondrial fractions from a cell line co-expressing C-terminally c-Myc-tagged ATOM14 and C-terminally HA-tagged ATOM40. Immunoblots containing 5% input (In) and 100% eluate (IP) were probed with an anti-ATOM14 antiserum that recognizes both the tagged als well as the endogenous ATOM14. HA-tagged ATOM40 and VDAC serve as a controls. (C) Immunoblots of the total (T), pellet (P) and supernatant (S) fractions of carbonate extracted mitochondria isolated from cells expressing C-terminally c-Myc-tagged ATOM14 were analyzed by anti-c-Myc antiserum. VDAC and cytochrome C (Cyt C) serve as marker for an integral and peripheral membrane protein, respectively. (D) Immunoblots of a protease protection assay using gradient-purified mitochondria isolated from cell lines expressing N- or C-terminally c-Myc-tagged ATOM14, respectively, analyzed by anti-c-Myc antiserum. The IMS protein Tim9 serves as a control.

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