1. Structural Basis for Protein Recognition by B30.2/SPRY Domains
Jae-Sung Woo, Hye-Young Suh, Sam-Yong Park, Byung-Ha Oh 
Molecular Cell 
Volume 24, Issue 6, Pages (December 2006)
DOI: /j.molcel
Copyright © 2006 Elsevier Inc. Terms and Conditions

2. Figure 1 Structures of GUSTAVUS
(A) B30.2/SPRY domain in complex with peptide I. Loop A to loop E (blue), extending from the two β sheets (green and orange), constitutes the peptide-binding site. Peptide I (yellow) is shown as a coil for the main chain and sticks for the side chains. The secondary structural elements of the domain are sequentially labeled. The domain organizations of GUSTAVUS and VASA are shown. The regions in green and yellow on the diagrams indicate the fragments of the proteins used for the structure determination.
(B) Structural superposition. The GUSTAVUS (residues 29–234)-peptide I complex is superposed on the GUSTAVUS (residues 29–253)-Elongin BC complex (PDB code 2FNJ), using the program SuperPose (Maiti et al., 2004). The B30.2/SPRY domain and peptide I are colored as in (A), and the BC box of GUSTAVUS is in magenta. The two superposed B30.2/SPRY domains are hardly discernible due to a very small structural difference between the two complexes.
Molecular Cell  , DOI: ( /j.molcel )
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3. Figure 2 Interaction of GUSTAVUS with Peptide I
(A) A detailed view. The B30.2/SPRY domain is shown as a ribbon drawing in the transparent surface. The residues involved in the intermolecular interactions between peptide I (yellow) and the domain (white) are represented as sticks. Labeled are loop A to loop E and the residues involved in the intermolecular interactions. Dotted lines indicate the intermolecular hydrogen bonds. A red sphere indicates a bound water molecule.
(B) Shape complementarity. The interaction of the protein (mesh model) with peptide I (CPK model) is superficial except for the pocket that accepts the side chains of Asn186, Asn187, and Asn188 (not visible). Ile185 caps the interaction between Asn186 and the pocket.
(C) ITC analysis. The interactions between the B30.2/SPRY domain of GUSTAVUS and the indicated VASA peptides were analyzed by ITC, and the measured KD values are shown.
Molecular Cell  , DOI: ( /j.molcel )
Copyright © 2006 Elsevier Inc. Terms and Conditions

4. Figure 3 Interaction of Par-4 Peptides with the SSB Family Members
(A) Sequence alignment. The red, pink, and gray columns indicate the residues similarly conserved in five, four, and three of the aligned proteins, respectively. The secondary structural elements of GUSTAVUS are shown at the top of the alignment. The blue lines and labels indicate loop A to loop E. The triangles indicate the peptide I-interacting residues of GUSTAVUS. Of these, the pocket-forming residues are indicated by the filled triangles.
(B) E-L-N-N-N-L sequence motif in Par-4. The motif is indicated by the green bar on the diagram for the primary structure of Par-4. “SAC” and “LZ” stand for the core domain causing selective apoptosis of cancer cells and the leucine zipper region, respectively. The VASA segment containing the D-I-N-N-N-N sequence is aligned with two Par-4 segments containing the E-L-N-N-N-L sequence and a Par-4 segment containing a N72A substitution.
(C) ITC analysis. The Par-4 peptides were titrated into the indicated protein solutions, and the measured KD values are summarized in the table. The SSB-2 (HSVG) mutant contains the His116-Ser117-Val118-Gly119 sequence (see text).
(D) (His)6-tag pull-down assay. Each of the wild-type and N72A mutant Par-4 (100 μg for each) was mixed with (His)6-tagged SSB-1 (50 μg) and Ni2+-chelating resin. After vigorous washing, the resin-bound proteins were analyzed by denaturing gel electrophoresis.
Molecular Cell  , DOI: ( /j.molcel )
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5. Figure 4
Presence of a Preformed Pocket in PSD and a Putative Pocket in Other B30.2/SPRY Domains
(A) A preformed pocket on surface A of PSD. The ribbon drawing and surface representation of PSD (PDB code 2FBE) are in the same orientation with GUSTAVUS. The presence of a pocket in PSD (middle) and GUSTAVUS (right) is highlighted by coloring the rim residues. The red meshes on PSD indicate the mapping of the FMF-causing mutations in Pyrin (red letters). The corresponding residues of PSD are in parentheses. “del” stands for deletional mutation.
(B) High sequence homology between PSD and 17 TRIM family proteins. Aligned are the sequences of the B30.2/SPRY domains of PSD and 17 TRIM family proteins (see text). The variable regions are indicated at the bottom of the alignment. Cyan and pink columns indicate the residues similarly conserved in more than 14 and 10 of the aligned proteins, respectively. Magenta and red boxes indicate the pocket-forming residues of PSD and the residues of Pyrin mutated in the FMF patients, respectively.
(C) Location of the conserved residues. On the ribbon drawing of the PSD structure, the conserved residues are shown as sticks in the same color as in (B). The circle indicates the location of the pocket on surface A.
Molecular Cell  , DOI: ( /j.molcel )
Copyright © 2006 Elsevier Inc. Terms and Conditions