The Arabidopsis TAG1 transposase has an N-terminal zinc finger DNA bindingdomain that recognizes distinct subterminal motifs

The in vitro DNA binding activity of the Arabidopsis Tag1 transposase (TAG1 ) was characterized to determine the mechanism of DNA recognition. In addit ion to terminal inverted repeats, the Tag1 element contains four different subterminal repeats that flank a transcribed region encoding a 729-amino ac id protein. A single site-specific DNA binding domain is located near the N terminus of TAG1, between residues 21 and 133. This domain binds specifica lly to the AAACCC and TGACCC subterminal repeats, found near the 5' and 3' ends of the element, respectively. The ACCC sequence within these repeats i s critical for recognition because mutations at positions 3, 5, and 6 aboli shed binding, yet the first two bases also are important because substituti ons at these positions decreased binding by up to 90%. Weak interaction als o occurs with the terminal inverted repeats, but no binding was observed to the other two 3' subterminal repeat regions. Sequence analysis of the TAG1 DNA binding domain revealed a C2HC zinc finger motif. Tests for metal depe ndence showed that DNA binding activity was inhibited by divalent metal che lators and greatly enhanced by zinc. Furthermore, mutation of each cysteine residue predicted to be a metal ligand in the C2HC motif abolished DNA bin ding. Together, these data show that the DNA binding domain of TAG1 specifi cally binds to distinct subterminal repeats and contains a zinc finger.