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A homeodomain is a highly conserved protein domain (first identified in Antp and Ubx proteins) containing 60 amino acids that fold into three conserved DNA-binding helics (indicating a role in transcriptional reglation). Many homeodomain proteins have been shown in vitro to bind to regulatory regions of genes known to regulate by genetic criteria. The DNA sequence encoding a homeodomain is a homeobox.

Specific and highly conserved residues in the homeodomain's third helix (helix 3) bind the major groove of DNA, meaning all homeodomain proteins bind the same DNA sequence in vitro. Specific homeodomain proteins have different regulatory properties via interaction with different co-factors; the co-factor and the homeodomain protein bind a specific DNA sequence together, but are unable to bind it individually.