Existing at the heart of cellular energy metabolism, the mitochondrion is uniquely positioned to have a major impact on human disease processes. Examples of mitochondrial impact on human pathology abound and include etiologies ranging from inborn errors of metabolism to the site of activity of a variety of toxic compounds. In this review, the unique aspects of the mechanisms related to the mitochondrial proteome are discussed along with an overview of the literature related to mitochondrial proteomic exploration. The review includes discussion of potential areas for exploration and advantages of applying proteomic techniques to the study of mitochondria.