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. 2008 Oct 24;283(43):29126–29134. doi: 10.1074/jbc.M803028200

FIGURE 2.

FIGURE 2.

Subcellular localization of cytosol- (A), matrix- (B), and IMS-rxYFP (C). WT yeast cells (BY4741) expressing cytosol-, matrix-, or IMS-rxYFP were grown to mid-log phase in SC galactose media. Cells were lysed and fractionated, and fractions were analyzed by SDS-PAGE and immunoblotting using antibodies directed against rxYFP, PGK1 (cytosol marker), Pos5 or Mas2 (mitochondrial matrix markers), or Cyb2 (mitochondrial IMS marker). In the left panel of A–C, 75 μg of total cell protein (Total) was fractionated into post-mitochondrial supernatant (PMS) and mitochondria (Mito), and the entire amount of each fraction was analyzed. In the right panel of B, mitochondria (15 μg of protein) from cells expressing matrix-rxYFP were further fractionated into IMS and mitoplast components, and the entire amount of each fraction was analyzed. In the right panel of C, mitochondria (15 μg of protein) from cells expressing IMS-rxYFP were fractionated as in B. Mitoplast and IMS fractions were further treated with proteinase K and/or Triton X-100 as indicated. rxYFP-expressing plasmids utilized include: pJH208 (cytosol), pLD207 (matrix), and pJH200 (IMS).