We present the structure and nucleotide sequence of a gene encoding the human epidermal 67-kDa keratin. Three genomic clones were isolated from a lambda Charon 4A human genomic library by hybridization to a specific cDNA probe. One clone of 12.3 kilobase pairs was shown by R-loop, DNA sequence, and primer-extension analyses to encode an entire gene of about 6.25 kilobase pairs. Of eight identified introns, seven are located within the region that encodes the central coiled-coil alpha-helical domain of the protein. Except for one intron located at the end of the region encoding this domain, these do not delineate apparent structural subdomains. The positions of five of the introns exactly coincide with the positions of introns previously reported in the hamster gene for the intermediate filament protein vimentin [Quax, W., Egberts, W.V., Hendricks, W., Quax-Jeuken, Y. & Bloemandal, H. (1983) Cell 35, 215-233]. These findings suggest that the human 67-kDa keratin and vimentin genes arose from a common ancestral gene.