THAP proteins target specific DNA sites through bipartite recognition of adjacent major and minor grooves.
THAP-family C(2)CH zinc-coordinating DNA-binding proteins function in diverse eukaryotic cellular processes, such as transposition, transcriptional repression, stem-cell pluripotency, angiogenesis and neurological function. To determine the molecular basis for sequence-specific DNA recognition by THAP proteins, we solved the crystal structure of the Drosophila melanogaster P element transposase THAP domain (DmTHAP) in complex with a natural 10-base-pair site. In contrast to C(2)H(2) zinc fingers, DmTHAP docks a conserved beta-sheet into the major groove and a basic C-terminal loop into the adjacent minor groove. We confirmed specific protein-DNA interactions by mutagenesis and DNA-binding assays. Sequence analysis of natural and in vitro-selected binding sites suggests that several THAPs (DmTHAP and human THAP1 and THAP9) recognize a bipartite TXXGGGX(A/T) consensus motif; homology suggests THAP proteins bind DNA through a bipartite interaction. These findings reveal the conserved mechanisms by which THAP-family proteins engage specific chromosomal target elements.
Sabogal
Alex
A
Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA.
Lyubimov
Artem Y
AY
Corn
Jacob E
JE
Berger
James M
JM
Rio
Donald C
DC
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002850:Chromatography, Gel; D003460:Crystallization; D004247:DNA / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster / Q000201:enzymology*; D024202:Electrophoretic Mobility Shift Assay; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary / Q000235:genetics*; D017422:Sequence Analysis, DNA; D019895:Transposases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D017931:DNA Primers; D004268:DNA-Binding Proteins; D004247:DNA; C578085:THAP9 protein, human; D019895:Transposases