The gene encoding cytosolic adenylate kinase (AK1) was isolated from a chicken genomic DNA library by using its cDNA as a hybridization probe. The chicken AK1 gene spanned about 6 kilobase pairs and consisted of 7 exons. Analyses of the 5'-flanking region sequence and S1 nuclease mapping revealed that the transcription initiation site is located at 84 base pairs upstream from the ATG initiation codon. The TATA box and the putative CAT box were located 29 base pairs and 97 base pairs upstream from the transcription initiation site, respectively. A total of 7 GC boxes were found in the 5'-flanking region, the exon 1, and the intron 1. The GC boxes were surrounded by the sequences with extremely high G+C contents. When projected on the three-dimensional structure of the AK1 protein molecule, introns fell either between or near the ends of alpha-helices and beta-strands, and most of the coding exons encoded at least one alpha-helix and one beta-strand. The dot matrix plot analysis between chicken AK1 and bovine mitochondrial adenylate kinase (AK2) suggested that the AK1 gene might have evolved from the AK2 gene by deletion of one or more exon(s).