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. 2019 Mar 12;8:e44771. doi: 10.7554/eLife.44771

Figure 3. Structure and interactions of XPB.

(A) Bottom lobe of TFIIH. XPB RecA1/2 teal, DRD blue, NTD dark blue, p52 yellow, p8 green, p44 NTE red. (B) Superposition of the XPB NTD and the p52 clutch domain. (C) Mapping of mutations on the XPB NTD and the p52 clutch domain; mutated regions are color-coded or shown as spheres (see text for details). (D) The combined interactions of the p52 clutch, the p8-p52 CTD dimer, and the XPB NTD with XPB RecA2 may restrict the conformational flexibility of XPB RecA2 to optimize XPB activity. (E) An extension of the DRD (blue) contacts XPD (green). The sequence for which formation of an α-helix is predicted (Kelley et al., 2015) is indicated. (F) The DRD extension overlaps with the substrate-binding site on XPD RecA2. Substrate DNA modeled from PDB ID 5HW8 (Constantinescu-Aruxandei et al., 2016).

Figure 3.

Figure 3—figure supplement 1. Domain organization of XPB and sequence alignment of the N-terminal regions of XPB or XPB-like enzymes from eukaryotes, archaea, and bacteria.

Figure 3—figure supplement 1.

(A) Domain organization of human XPB, color-coded as in Figure 3A. (B) Sequence alignments. Sequences used (species and UNIPROT or GENBANK accession code indicated): Human (Homo sapiens, P19447), mouse (Mus musculus, P49135), fruit fly (Drosophila melanogaster, Q02870), Choanoflagellate (Monosiga brevicollis, A9V0A0), fungus (Saccharomyces cerevisiae, Q00578), ciliate (Paramecium tetraurelia, XP_001448651.1), plant (Arabidopsis thaliana, Q38861), archaea (Ferroplasma, EQB73227.1, Thermoplasmatales, EQB71900.1), bacteria (Mycobacterium leprae, Q9CBE0) (Balasingham et al., 2012; Poterszman et al., 1997). Some archaeal XPBs, including the Archaeoglobus fulgidus homolog that served for structure determination (Fan et al., 2006), contain only the DRD, lacking the NTD/NTE, and were not included. Residues are numbered according to the human sequence. Identical residues are printed white on red background, similar residues in red on white background. Secondary structure elements, residues mutated in human disease (F99S, T119S), and the approx. 70-residue insertion in the DRD are indicated. Figure generated using the ESPript web server (Robert and Gouet, 2014).
Figure 3—figure supplement 2. Structure and interactions of the XPB DRD and NTD.

Figure 3—figure supplement 2.

(A) Interface between the XPB NTD and the p52-clutch domains. Contacts include several large hydrophobic residues, as well as a salt bridge between p52 R314 and XPB E115 (shown as sticks). Mutations of residues equivalent to R314 and E310 in D. melanogaster p52 lead to disease-like phenotypes (Fregoso et al., 2007). (B, C) Electrostatic surface potential of the interacting surfaces of the XPB NTD (B) and the p52 clutch domain (C) show charge complementarity (strongest complementary charge peak indicated by a dotted circle). (D) Front and back views of the domain organization of p52. Residues 1–130 (red) and 131–304 (orange) are not in contact with the XPB NTD. Residues 304–400 of p52 form the p52 clutch. Protein subunits or domains interacting with p52 are shown in grey. (E) Same as D), but p52 is colored using a gradient from blue (N-terminus) to red (C-terminus). (F) Sequence alignment of human and yeast p52 clutch and XPB NT domains highlights several highly conserved residues, including XPB T119, which is affected by a human disease mutation. Generated using the ESPript web server (Robert and Gouet, 2014). (G) Location of XPB T119, affected by the TTD mutation T119P, at a junction between a β-strand and an α-helix near the interface between XPB NTD, NTE, and RecA1 domains. (H) Superposition of the XPB NTD and the p52 clutch shows that a threonine at the position of XPB T119 also occurs in the p52 clutch. (I) The XP/CS mutation F99S affects a XPB NTD residue (orange) situated in a hydrophobic pocket formed by aliphatic residues (cyan) that are conserved across yeast and human XPB and p52 (F).