Supplementary Figure 5: YL1 ZID–H2A.Z–H2B is monomeric in solution, and the YL1 and SWC2 N-terminal regions (ZIDN) are required for proper binding to human H2A.Z but not to yeast HTZ1.
From: Molecular basis and specificity of H2A.Z–H2B recognition and deposition by the histone chaperone YL1
a) Size exclusion chromatography purification profile on a Superdex 200 16/60 (left) and analytical ultracentrifugation (AUC) data of the YL1–ZID–H2A.Z–H2B complex. This analysis shows that the YL1–ZID–H2A.Z–H2B complex is monomeric in solution.
b) Ribbon representation of the YL1–ZID–H2A.Z–H2B complex as observed in the structure (left; ZIDN colored violet in a position interacting with a symmetry-related H2A.Z–H2B dimer) and of the model of the YL1–ZID–H2A.Z–H2B complex (right; ZIDN colored green interacting with the same H2A.Z–H2B dimer than the rest of the YL1–ZID domain). The model was created by modeling the ZIDN onto the H2A.Z–H2B dimer recognized by the rest of the YL1–ZID and using the mode of binding observed when it binds to a symmetry-related H2A.Z–H2B dimer. The model was then refined to obtain correct stereochemistry. This procedure showed that only the position of YL1–ZID residues E24, E25 and E26 (colored blue), which are part of the hinge region between YL1–ZID αN1 and αN2 helices, need to be moved to adapt to the new ZIDN conformation.
c) Sequence of yeast SWC2–ZID from alignment in figure 2b. Stars below the alignment indicate the residues mutated in either mutant m1 (dark blue) or mutant m2 (light blue). Mutant SWC2-ΔN was made by deletion of the N-terminal region of SWC2 (deletion of amino acids 1–45), including the αN1 helix. Mutant SWC2-m1 (I63A L65A L66A F67A) targeted the H2A.Z αC binding region of YL1 and mutant SWC2-m2 (D73A D75A F76A) targeted the first DNA-binding region of SWC2.
d) His-pulldowns with yeast His–SWC2–ZID and HTZ1/HTB1 complexes using WT HTZ1 and the indicated SWC2 mutants. The interaction between SWC2 and HTZ1–HTB1 is strongly affected by the m1 and m2 mutations but not by deletion of the N-terminal region (ΔN).
e) GST-pulldowns with human GST–YL1–ZID (1–69) or GST–YL1–ZID–ΔN (25–69) and human H2A.Z–H2B or yeast HTZ1–HTB1 dimers. The interaction between human YL1 and histones is dependent of its N-terminal region (ZIDN) for proper binding to human H2A.Z–H2B but not to yeast HTZ1–HTB1.‘*’, degradation products.
f) His-pulldowns with yeast His–SWC2–ZID (1–101) or His–SWC2–ZID–ΔN (46–101) and human H2A.Z–H2B or yeast HTZ1–HTB1 dimers. The interaction between yeast SWC2 and histones is dependent of its N-terminal region (ZIDN) for proper binding to human H2A.Z–H2B but not to yeast HTZ1–HTB1.
