In the course of studies on mutations in human mitochondrial (mt) DNA, we have uncovered and sequenced four new nuclear pseudogenes corresponding to bp 2457-2657 of the mt 16S rDNA. The four genes and their homologies with human mtDNA are E2 (62.4%), K10 (74.4%), E1 (84.6%) and LE6 (93.2%). When these five pseudogene sequences and another previously reported pseudogene sequence are compared with each other, they display what appears to be an ordered series of steps from a hypothetical common ancestor. The sequence of the hypothetical ancestor closely resembles that found in a wide variety of present-day mammalian mt genomes. The pseudogene sequences suggest an evolutionary trail of mt mutation dominated by base pair transitions punctuated by integration into the nuclear genome. Once integrated into the nuclear genome, the pseudogenes appear to follow the distinctive nuclear mutational pathway in which GC to AT transitions predominate and CpG sequences are preferentially eliminated.