BESS domain is a protein binding domain that can interact each other or with other domains. In this study, 323 BESS domain containing (BESS-DC) proteins were identified in 3328 proteomes. These BESS-DC genes pertain to 41 species of five phyla, most of which are arthropod insects. A BESS domain contains two α-helixes linked by a coil or β-turn. Phylogenetic tree and architecture analysis show that the BESS domain seems to generate along with the DNA-binding MADF domain. Two hundred thirty three BESS-DC genes (71.1%) contain at least one MADF domain, while 59 genes (18.2%) had only the BESS domain. In addition to BESS and MADF domains, some of genes also contain other ligand binding domains, such as DAO, DUS and NAD_C. Nineteen genes (5.8%) are associated with other DNA binding domains, such as Myb and BED. The BESS-DC genes can be divided into 17 subfamilies, eight of which have more than one clade. In Bombyx mori, 12 BESS-DC genes that do not contain intron in the BESS domain region were localized to eight chromosomes. Real-time PCR results showed that most of the B. mori BESS-DC genes highly expressed from late larval stage to adult stage. The results of sequence comparison and evolution analyses suggest a hypothesis that the BESS-DC genes may play a role in central nervous system development, long term memory and metamorphosis of insects of different phyla.