No abstract

The Ets family of transcription factors, of which there are now about 35 members regulate gene expression during growth and development. They share a conserved domain of around 85 amino acids which binds as a monomer to the DNA sequence 5'-C/AGGAA/T-3'. We have determined the crystal structure of an ETS domain complexed with DNA, at 2.3-A resolution. The domain is similar to alpha + beta (winged) 'helix-turn-helix' proteins and interacts with a ten-base-pair region of duplex DNA which takes up a uniform curve of 8 degrees. The domain contacts the DNA by a novel loop-helix-loop architecture. Four of amino acids that directly interact with the DNA are highly conserved: two arginines from the recognition helix lying in the major groove, one lysine from the 'wing' that binds upstream of the core GGAA sequence, and another lysine, from the 'turn' of the 'helix-turn-helix' motif, which binds downstream and on the opposite strand.

When compared with the structure of the coat protein in the assembled virus, differences in orientation of residues 85 and 87 suggest conformational adjustment on binding RNA in the first step of viral assembly. The substitution at residue 82 may affect virus assembly by imposing conformational restriction on the loop that makes critical inter-subunit contacts in the capsid.

Transcription factors bind to target DNA sequences to regulate metabolic functions such as growth and differentiation. Typically, the molecular scaffold for DNA recognition is conserved within a given family of DNA-binding proteins. In some cases the similarity of these scaffolds suggests an evolutionary relationship between different families or comparison of scaffolds reveals a structural similarity that was obscured by sequence comparisons alone.A recently discovered family of regulatory proteins, the ets gene family, includes more than 45 members in a variety of organisms from Drosophila to humans (1, 2). These molecules play a role in normal development and have been implicated in malignant processes such as erythroid leukemia and Ewing's sarcoma. The DNA-binding domain of ets proteins is a conserved region (ETS domain) that is about 85 residues in length. Although ets proteins share a homologous sequence in the ETS domain, they differ in length and in the relative position of this domain. In some molecules, the ETS domain is found at the carboxyl terminus (e.g. PU.1 (3); ets-1 (4); ets-2 (5)), while in others the domain is located in the middle of the sequence (erg (6)), or in the amino-terminal region (elk-1 (7)). Flanking regions are thought to form other functional domains that influence protein-protein recognition or inhibitory domains that mask the DNA-binding site (8, 9).1 In ets-1, an ␣-helix that is located in an inhibitory domain immediately NH 2 -terminal to the ETS domain unfolds on DNA-binding (10). Regardless of the position of the ETS domain within the intact ets proteins, there is strong sequence homology in this conserved region.We have determined the crystal structure of the ETS domain of the PU.1 transcription factor complexed to DNA (11). The domain is similar to ␣ϩ␤ helix-turn-helix (HTH) 2 DNA-binding proteins and contacts a 10-base pair region of duplex DNA that is bent (8°) but uniformly curved without distinct kinks. The PU.1 domain assumes a tight globular structure with three ␣-helices and a four-stranded antiparallel ␤-sheet enclosing a hydrophobic core. The topology of the domain is similar to the structures of other ets family proteins fli-1 (12), murine ets-1 (13), and human ets-1 (14) determined in solution by NMR. The common molecular scaffold is similar to DNA-binding proteins such as CAP (15) and resembles "winged"-HTH proteins including HNF-3␥ (16). ETS domains bind as a monomer to the core sequence 5Ј-(C/A)GGA(A/T)-3Ј.The PU.1 domain contacts DNA from three sites: the recognition helix (␣3) interacts with the GGAA core sequence in the major groove, while contacts with the phosphate backbone on either side of this site are made in the minor groove by two loops. Therefore, the PU.1 ETS domain binds DNA by a loophelix-loop motif. One loop is formed between ␤-strands 3 and 4 (a "wing") and the other is a loop in the position of the turn in the HTH motif (␣2-turn-␣3). The protein-DNA contacts stabilize a uniform bending of the duplex DNA that likely is due to phosphate neutra...