The multigene family of chromosomal protein HMG-17 is the largest known human retropseudogene family. A functional gene was identified and isolated by screening cDNA-selected genomic clones with a set of 5 oligonucleotides whose sequence corresponded to regions in which the sequence of the retropseudogenes differed from that of the cDNA and which did not span previously identified exon/intron junctions. A 7195 bp genomic fragment containing 6 exons, ranging in size from 30 to 817 bp, two of which encode the entire DNA binding domain of the protein, was sequenced. The gene has features which are typical to "housekeeping" genes and is characterized by a very high content of G + C residues in a 1.4 kb fragment starting 500 bp from the cap site and by an "HTF" island in the 5' region. Transcriptional regulatory signals, exon/intraon boundaries and features characteristic of "housekeeping" genes are evolutionary conserved between the human and chicken genes. The HMG-17 gene was localized to human chromosome 1p12-34. RFLP's useful for further mapping were detected. The experimental evidence presented leads to the assumption that the gene characterized is the only functional human HMG-17 gene.