Subunit 6 of the yeast cytochrome bc1 complex contains a 25 amino acid presequence that is not present in the mature form of the protein in the bc1 complex. The presequence of subunit 6 is atypical of presequences responsible for targeting proteins to mitochondria. Whereas mitochondrial targeting sequences rarely contain acidic residues and typically contain basic residues that can potentially form an amphiphilic structure, the presequence of subunit 6 contains only one basic amino acid and is enriched in acidic amino acids. If the 25 amino acid presequence is deleted, subunit 6 is imported into mitochondria and assembled into the cytochrome bc1 complex and the activity of the bc1 complex is identical to that from a wild-type yeast strain. However, if the C-terminal 45 amino acids are truncated from the protein, subunit 6 is not present in the mitochondria and the activity of the bc1 complex is diminished by half, identical to that of the bc1 complex from a yeast strain in which the QCR6 gene is deleted. These results indicate that the presequence of subunit 6 is not required for targeting to mitochondria or assembly of the subunit into the bc1 complex and that information necessary for targeting and import into mitochondria may be present in the C-terminus of the protein.