Structure of the DNA-Bound T-Box Domain of Human TBX3, a Transcription Factor Responsible for Ulnar-Mammary Syndrome  Miquel Coll, Jonathan G Seidman, Christoph W Müller  Structure  Volume 10, Issue 3, Pages 343-356 (March 2002) DOI: 10.1016/S0969-2126(02)00722-0

Figure 1 Multiple Sequence Alignment and DNA Binding Sites of T-Box Transcription Factors (A) Sequence alignment of the T-box domains of human TBX3 (SwissProt accession number O15119), TBX1 (O43435), TBX2 (Q13207), TBX4 (P57082), TBX5 (Q99593), TBX6 (O95947), and the T protein from mouse (P20293) and Xenopus laevis (Xbra; P24781). Sequence numbering and secondary structure elements for TBX3 and Xbra are given above and below the sequences, respectively. For the other T-box domain sequences, the numbering of the first residue of each line is provided. Identical residues are depicted on a light yellow background. TBX3 and Xbra residues involved in interaction with the DNA or with the second monomer are depicted on blue and magenta backgrounds, respectively. Point mutations observed in UMS (TBX3) and HOS (TBX5) patients are marked in green. (B) DNA duplex present in the TBX3/DNA complex crystals containing the palindromic DNA binding site aligned with half-sites found in promoters of target genes of T-box transcription factors. The 20 base pair palindromic binding sequence is drawn in a box. The dyad is indicated by a black circle. Target sequences for mouse and human FGF, eFGF, Bix4, TRP-1, and POMC genes are denoted in the 5′–3′ direction. One half-site of the original palindromic binding site and bases in target sites conserved with respect to the original half-site are depicted in yellow. For the eFGF-proximal and Bix4-distal sites, no direct binding of T-box factors has been demonstrated [24]. Structure 2002 10, 343-356DOI: (10.1016/S0969-2126(02)00722-0)

Figure 2 Structure of the TBX3/DNA Complex (A) Ribbon diagram of a human TBX3 monomer bound to its DNA target site. Secondary structure elements are labeled, and helices, strands, and loops are depicted in turquoise, red, and gray, respectively. Figures 2A, 3A, 3B, and 5 were produced with the program SETOR [40]. (B) Stereodiagram of the superposition of the Cα backbone of the TBX3 monomer with Xbra (thick and thin lines, respectively). (C) Final 2Fo−Fc electron density at 1.5σ showing the recognition of G:C base pair 5 by Arg130 and Phe283 in the major and minor groove, respectively. Structure 2002 10, 343-356DOI: (10.1016/S0969-2126(02)00722-0)

Figure 3 DNA Recognition by Human TBX3 (A) Interactions between one TBX3 monomer and one half-site. The view corresponds to Figure 2A. Depicted are all TBX3 residues involved in hydrophobic and polar interactions with one half-site of the palindrome. (B) Interaction of Arg284 with backbone carbonyls of Ile274 and Asn277 connecting helix 310C with helix α3. (C) Schematic diagram of the interactions between one monomer and one DNA half-site. Polar and hydrophobic interactions are indicated by arrows and dashed lines, respectively. Residues involved in polar interactions and residues contacting the DNA only through hydrophobic interactions are drawn on red and blue labels, respectively. Structure 2002 10, 343-356DOI: (10.1016/S0969-2126(02)00722-0)

Figure 4 Minor Groove Width and Water Structure (A) Schematic diagram of minor groove width and the water molecules present in the minor groove. Depending on minor groove width, phenylalanine side chains Phe279 and Phe283 (depicted as red squares) and the backbone carbonyl of residue 279 (yellow label) displace one or two water molecules (green spheres). Distances between phosphorous atoms across the minor groove are given in Angstroem. (B) Minor and major groove width of TBX3 (thick lines) and Xbra (thin lines) per base step. Groove widths were calculated according to [43]. Major groove width for base step k between strands I and II: (P(I)k+2)−(P(II)k−2); minor groove width for base step k: 1/2((P(I)k+1−P(II)k−2)+(P(I)k+2−P(II)k−1)). The P(I)k+1−P(II)k−2, P(I)k+2−P(II)k−1 distances across the minor groove are given in (A). Major and minor groove widths for B-form DNA of 17.4 and 10.8 Å, respectively are indicated by horizontal lines. These values are based on crystal structures of oligonucleotides [39]. Structure 2002 10, 343-356DOI: (10.1016/S0969-2126(02)00722-0)

Figure 5 Quaternary Structures of TBX3 and Xbra T-Box Domains Bound to DNA (A) Ribbon diagram of the two TBX3 T-box domains bound to the palindromic DNA duplex. View perpendicular to the DNA axis and the noncrystallographic dyad. The two monomers are depicted in light gray and black. The two arrows indicate the rotations of both monomers in opposite directions with respect to the two monomers in the Xbra/DNA complex. The rotation axis runs at an angle of about 45° through the points of contact formed by Phe279 and Phe283, marked by asterisks in both monomers. (B) Stereodiagram of the two TBX3 monomers bound to palindromic DNA superimposed with the Xbra/DNA complex. Human TBX3 and Xbra are depicted in magenta and green, respectively. Only the DNA duplex of the TBX3/DNA complex is depicted. In both complexes, the central 12 DNA phosphates of both strands were superimposed (rms24 P atoms = 0.95 Å). View along the dyad of the complex. The direction of the rotational axis is depicted. (C) View along the DNA axis. Structure 2002 10, 343-356DOI: (10.1016/S0969-2126(02)00722-0)

Figure 6 Monomer/Monomer Contacts in the Xbra/DNA and TBX3/DNA Complexes (A) Stereodiagram of the monomer/monomer contacts observed in the Xbra/DNA complex. (A) and (B) were produced with the program MOLSCRIPT [42]. With respect to Figure 5B, the model was rotated clockwise by about 45°. (B) Monomer/monomer contacts observed in the human TBX3/DNA complex. Structure 2002 10, 343-356DOI: (10.1016/S0969-2126(02)00722-0)