Fig. 2. Abf2 protein interactions in mitochondria. (A) Identification of mitochondrial Abf2-interacting proteins. Purified mitochondria from yeast strains expressing Abf2 and Abf2-13Myc were crosslinked with DSP. Mitochondrial lysates were incubated with DNaseI to remove mtDNA, and Abf2 protein was immunoprecipitated with anti-Myc antibody. Protein samples were compared on silver-stained gels. Protein bands that were unique to the Abf2-13Myc sample were further examined. (B) Silver-stained 4%-20% gradient SDS-PAGE gel of the protein samples immunoprecipitated with an anti-Myc antibody from the mitochondria isolated from MCC109 cells expressing the wild-type Abf2 protein and the Abf2-13Myc fusion protein. Where indicated, mitochondria were incubated with the protein crosslinker DSP prior to incubation with the antibody. a, Kgd1; b, Aco1; c, Ald4; d, Abf2-13Myc; e, Idh2; f, Idh1. (C) Southern blot confirming that the DNA was degraded after incubation with DNAseI in samples used for coimmunoprecipitation in B. DNA was isolated from 5 µg of purified mitochondria and from an aliquot of DNAseI-treated lysate that contained 50 µg of proteins. MtDNA was probed using a PCR-amplified region of the mitochondrial COX2 gene.