LOCALIZATION OF BOUND WATER IN THE SOLUTION STRUCTURE OF A COMPLEX OFTHE ERYTHROID TRANSCRIPTION FACTOR GATA-1 WITH DNA

Background: The erythroid specific transcription factor GATA-1 is resp onsible for the regulation of transcription of erythroid-expressed gen es and is an essential component required for the generation of the er ythroid lineage. GATA-1 binds specifically as a monomer to the asymmet ric consensus target sequence (T/A)GATA-(A/G) found in the cis-regulat ory elements of all globin genes and most other erythroid specific gen es that have been examined. We have previously determined the solution structure of the complex of the zinc-containing DNA-binding domain of chicken GATA-1 with its cognate DNA target site by multidimensional h eteronuclear NMR. From previous studies of complexes between proteins and DNA, water appears to play an important role in DNA-protein recogn ition by mediating bridging hydrogen bonds between functional groups o n the protein and DNA bases. Solvation free energy calculations, howev er, suggest that hydrophobic interactions should exclude water from pa rts of the GATA-1:DNA interface. Results: Using water-selective two-di mensional heteronuclear magnetic resonance spectroscopy, we have ident ified the location of bound water molecules in the specific complex of chicken GATA-1 with DNA. A number of water molecules could be detecte d between the protein and the phosphate backbone, as well as at the so lvent exposed surface of the protein. However, no water molecules coul d be observed at the interface of the protein with the bases of the DN A. With only one exception, the bound water molecules have a residency time > 200-300 ps.Conclusions: Unlike other protein-DNA complexes, th e majority of specific interactions between GATA-1 and the DNA bases i n the major groove are hydrophobic in nature. The exclusion of water f rom the protein-DNA base interface in the major groove supports the vi ew that the specific binding energy is indeed dominated by hydrophobic effects.