1. Volume 28, Issue 6, Pages 1093-1101 (December 2007)
Crystal Structure of Human XLF: A Twist in Nonhomologous DNA End-Joining 
Sara N. Andres, Mauro Modesti, Chun J. Tsai, Gilbert Chu, Murray S. Junop 
Molecular Cell 
Volume 28, Issue 6, Pages (December 2007)
DOI: /j.molcel
Copyright © 2007 Elsevier Inc. Terms and Conditions

2. Figure 1
Structure and Sequence Conservation of the N-Terminal Region of Human XLF
(A) Sequence alignment of XLF homologs. Conserved residues are colored as follows: hydrophobic, yellow; negative charge, red; positive charge, blue; proline and glycine, brown; threonine and serine, green; cysteine, light blue; and glutamine and asparagine, purple. Regions of highly conserved residues are underlined in green, orange, red, and black. For clarity, XLF residues 249–282 from S. cerevisiae are not shown in the alignment.
(B) Stereo image of a single XLF1–224. β strand and α helix are in red and blue, respectively.
(C) XLF dimer observed in crystal asymmetric unit. Chains A and B are shown in teal and yellow. Conserved patches are labeled and colored to correlate with (A).
Molecular Cell  , DOI: ( /j.molcel )
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3. Figure 2 Functional Analysis of XLF1–224
(A) Stimulation of noncohesive end-joining requires the C terminus of XLF.
(B) DNA binding of XLF requires the C-terminal 75 amino acids. XLF1–224 and/or XRCC4 was tested by mobility gel shift using increasing amounts of protein (2-fold increase starting at 4 pmol in lane 2) and 100 ng of linearized dsDNA (2.6 kbp).
(C and D) Protein-protein interactions with XLF1–224, XRCC4, and LigaseIV654–911. Native PAGE (C). Bands from lanes 4, 5, and 7 in (C) were cut out and resolved by SDS-PAGE (D).
Molecular Cell  , DOI: ( /j.molcel )
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4. Figure 3 Structure-Based Comparison of Human XLF1-224
(A) Structure-based sequence alignment of XLF1–224 and XRCC41–211. Conserved residues are colored according to the designation in Figure 1. Based on the S. cerevisiae structure of Lif1-Dnl4 (tandem BRCT), triangles indicate putative residues of XRCC4 that mediate LigaseIV interaction. Reflecting asymmetric interactions in Lif1-Dnl4, triangles are colored either black or gray for interaction with subunits A or B of XRCC4. XRCC4 and XLF mutations are indicated by triangles in blue and orange, respectively.
(B) Overlay of XLF1–224 (orange) and XRCC41–201 (blue) bound to LigaseIV755–782 (yellow). The helical tail region of XLF (residues 128–171) was used to structurally align the corresponding region of XRCC4. The structure of XRCC4 bound to LigaseIV (PDB 1Z56) is reported in Dore et al. (2006).
(C) Overlay of N-terminal head domains of XLF1–152 and XRCC41–142, in orange and blue, respectively. Arrow indicates a 45° difference in the trajectory of XLF and XRCC4 C-terminal tail domains.
(D) Stereo image of XRCC4-XLF interface suggested by mutational analysis. XLF-A and XLF-B are contributed from separate dimers. Mutations inhibiting XRCC4-XLF interaction are circled in gray.
(E) Model for assembly of an XRCC4-XLF-LigaseIV filament.
Molecular Cell  , DOI: ( /j.molcel )
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5. Figure 4 Mutational Analysis of XRCC4 and XLF Binding Surfaces
XRCC4 and XLF mutants either bound (+) or did not bind (−) DNA, LigaseIV654–911, XLF, or XRCC4. Stimulation of end-joining, illustrated as a bar graph, was measured from 0- to 150-fold for the joining of an EcoRV-KpnI-digested DNA substrate.
Molecular Cell  , DOI: ( /j.molcel )
Copyright © 2007 Elsevier Inc. Terms and Conditions