Solution structure of the ETS domain from murine Ets-1: a winged helix-turn-helix DNA binding motif.

Abstract

Ets‐1 is the prototypic member of the ets family of transcription factors. This family is characterized by the conserved ETS domain that mediates specific DNA binding. Using NMR methods, we have determined the structure of a fragment of murine Ets‐1 composed of the 85 residue ETS domain and a 25 amino acid extension that ends at its native C‐terminus. The ETS domain folds into a helix‐turn‐helix motif on a four‐stranded anti‐parallel beta‐sheet scaffold. This structure places Ets‐1 in the winged helix‐turn‐helix (wHTH) family of DNA binding proteins and provides a model for interpreting the sequence conservation of the ETS domain and the specific interaction of Ets‐1 with DNA. The C‐terminal sequence of Ets‐1, which is mutated in the v‐Ets oncoprotein, forms an alpha‐helix that packs anti‐parallel to the N‐terminal helix of the ETS domain. In this position, the C‐terminal helix is poised to interact directly with an N‐terminal inhibitory region in Ets‐1 as well as the wHTH motif. This explains structurally the concerted role of residues flanking the ETS domain in the intramolecular inhibition of Ets‐1 DNA binding.

Structural coupling of the inhibitory regions flanking the ETS domain of murine Ets‐1

“…8). Helices HI, H2, H3, and H4 show varying degrees of amphipathicity, consistent with their packing arrangement in the intact protein (Donaldson et al, 1996).…”

“…Two of the helices, helix HI1 and helix HI2, are located in the amino-terminal inhibitory region, helices (Hl-H3) comprise the DNA-binding wHTH ETS domain, and helix H4 is situated within the carboxy-terminal inhibition region. To maintain a consistent nomenclature, helices Hl-H4 correspond to those previously identified in Ets-lAN331 (Donaldson et a!., 1994(Donaldson et a!., , 1996, whereas HI1 and HI2 designate the two helices identified in the amino-terminal inhibitory region. Helix HI1 extends from Thr 303 to Asp 310 and helix HI2 -.…”

“…3). At the core of Ets-lAN280 is the previously identified wHTH ETS domain (Donaldson et al, 1994(Donaldson et al, , 1996Liang et al, 1994aLiang et al, , 1994b. Flanking the ETS domain is helix H4 in the carboxy-terminal inhibitory region and two helices, HI1 and HI2, in the amino-terminal inhibitory region.…”

“…The 3D structure of this protein has been determined by NMR methods ( Fig. 6; Donaldson et al, 1994Donaldson et al, , 1996. Ets-lAN331 lacks the aminoterminal inhibitory sequence and therefore binds to specific Ets-1 recognition sequences with approximately 15-fold higher affinity than either Ets-1AN280 or native Ets-1 (Jonsen et al, 1996;Petersen et al, 1995).…”

Molecular phylogeny of the ETS gene family

“…Despite various claims not substantiated by structural analysis, sequence comparisons have not revealed any significant sequence identities of the ETS domain to other well characterized DNA binding motifs such as the homeodomain, the zinc ®ngers or the helix ± turn ± helix. Nevertheless, the resolution of the three dimensional structure of the ETS domain of several Ets protein family members have revealed a wingedhelix ± turn ± helix (WHTH) structure, showing a striking structural similarity with the DNA binding motifs of the Escherichia coli catabolite gene activator protein, the HNF3/fork head family or the HSF (Heat Shock Factors) which all harbor a classical helix ± turn ± helix structure (Donalson et al, 1994(Donalson et al, , 1996Liang et al, 1994;Kodandapani et al, 1996). The ets gene family may thus be distantly linked at the evolutionary level to a large and extremely ancient winged-helix ± turn ± helix superfamily of transcriptional regulators.…”