A pumpkin cDNA for protein that binds to silencer of ascorbate oxidase gene was isolated by southwestern screening. The cDNA encodes a polypeptide of 380 amino acid residues. The protein, named AOBP (Ascorbate Oxidase gene Binding Protein), was shown to have a novel DNA-binding domain that has a significant resemblance to those of zinc fingers in steroid hormone receptors and GATA1. The novel DNA-binding domain, that is composed of 52 amino acid residues, has first been identified in AOBP and MNB1a, a maize DNA-binding protein. Furthermore, the DNA-binding domain, named Dof (DNA-binding with one finger) domain, has been found in DNA-binding proteins such as Arabidopsis OBP1, tobacco NtBBF1 and maize PBF1. Recent survey of databases shows that Dof domain occurs in at least ten proteins of Arabidopsis. Thus, Dof domain is highly conserved in DNA-binding proteins of higher plants. The N-terminal region of Dof domain may function as a Cys2/Cys2 zinc finger. Four Cys residues are used for coordinating to zinc ion and therefore are essential for DNA-binding. Two aromatic residues found C-terminal to the Cys residues may be positioned on the opposite side of the DNA and fix the structure so that the putative recognition helix may interact with DNA, as suggested in zinc fingers of steroid hormone receptors and GATA1. On the other hand, the putative loop region of Dof domain is much longer than those of steroid hormone receptors and GATA1 and has one more Cys residue near center of the loop. The Cys residue may negatively regulate DNA binding. Furthermore, Dof domain exists in plant proteins as a single unit although steroid hormone receptors and GATA1 have two zinc finger units. Thus, it is proposed that the Dof domain may be an unique zinc finger that is widely utilized for DNA-binding in plant kingdom.