Additional file 1:

Analysis of All Dimer Pairs Found in the IpgC^10-155 Asymmetric Unit. All nine IpgC^10-155 dimer pairs were superimposed by Local-Global Alignment to examine their overall similarity to one another. (A) Two orthogonal stereoscopic views of each dimer pair superimposed. A legend describing the identity of each protein chain in the corresponding PDB entry (accession code 3KS2) is shown underneath. (B) Quantitative analysis of all dimer superpositions from panel A presented in Table format.

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Additional file 2:

Comparison of Alternative Dimer Assemblies in the IpgC^10-155 Crystal. Potential contacts between either non-crystallographic or crystallographic symmetry-related IpgC^10-155 chains were evaluated using the EBI Protein Interfaces Surfaces and Assemblies (PISA) server[23]. (A) Two views of the rotationally-symmetric "head-to-head" dimer (as shown in Figure 1B, C) that is found in nine copies within the IpgC^10-155 asymmetric unit. On average, this arrangement buries 1262.5 Ų of surface area upon formation. (B) Two views of a rotationally-symmetric "tail-to-tail" dimer found in nine copies following application of crystallographic symmetry operators. On average, this arrangement buries 756.5 Ų of surface area upon formation. For the sake of clarity, the relative orientation of the orange colored IpgC^10-155 chain is identical between panels A and B.

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Additional file 3:

Comparison of IpgC^10-155 Dimers to Head-to-Head Dimer Assemblies Present in IpgC^1-151 Crystals. Both the free (3GYZ) and IpaB peptide-bound (3GZ1) structures of IpgC^1-151 were examined for potential "head-to-head" dimerization contacts similar to those observed in the IpgC^10-155 structure presented in Figure 1. (A) Two orthogonal views of a putative dimer (magenta and cyan) from 3GZ1 superimposed with a prototypic dimer of IpgC^10-155(blue and orange). This dimer is generated by applying the crystallographic symmetry operator x,-y+1,-z, and buries 666.9 Ų of surface area upon formation. (B) Two orthogonal views of a putative dimer from 3GYZ superimposed with the IpgC^10-155 dimer; all chains are colored as in panel A. This dimer is generated by applying the crystallographic symmetry operator x-y,-y,-z+2/3, and buries 950.4 Ų of surface area upon formation. (C) Two identical views of the IpgC^10-155 dimer (left) and the symmetry-generated dimer from 3GYZ shown in panel B (right). The set of aromatic residues described in Figure 3 are colored magenta and blue in the left and right panels, respectively.

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Additional file 4:

Magnified View of the Interface between the Amino-terminal Region and Helix α5 in an IpgC^10-155 Dimer. The individual monomers that comprise the IpgC^10-155 dimer are depicted as blue and orange cartoons, while the residues contributing to buried surface area (ball-and-stick) in this region are colored in magenta and cyan, respectively. Residues Ala⁹⁴ and Val⁹⁵, which are critical for dimerization in IpgC^1-151 [8], are colored yellow for reference. (A) Rendering of the IpgC^10-155 dimer from a viewing plane opposite that of Figure 1B, C. (B) Rotated view of panel A such that the axis of helix α5 is orthogonal to the plane of the page. Note the packing of the amino-terminal residues (magenta) against the sidechains of α5 (cyan and yellow). (C) Similar to panel B, but the viewing plane has been rotated 90° with respect to the page. The zigzag nature of the packing between the magenta-colored amino-terminus and the opposing monomer is evident in this image.

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Additional file 5:

IpgC Double Mutants Exist as Dimers in Solution. Description: Samples of purified IpgC (5 mg/mL) proteins were injected onto an analytical gel-filtration column and the elution profiles were compared to a series of known standards to derive an estimation of protein molecular weight (see Additional File 6, Figure S6). The sample identities are IpgC^10-155 (red), IpgC^10-155 Ala⁹⁴Glu/Val⁹⁵Gln (cyan), and IpgC^1-151 Ala⁹⁴Glu/Val⁹⁵Gln (purple). The standard mixture is shown as a black dashed line. Aside from the standard injection, all curves were normalized to a maximum peak height of 100 mAU for clarity.

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Additional file 6:

Calibration of the Analytical Size Exclusion Chromatography Column. Size exclusion standard curve where observed molecular weight is plotted as a function of elution volume. Calibration points (black diamonds) correspond to the following standard proteins: β-amylase (200 kDa), dehydrogenase (150 kDa), serum albumin (66 kDa), ovalbumin (43 kDa), carbonic anhydrase (29 kDa) and cytochrome c (12.4 kDa). Estimations of molecular weight were determined using the relationship MW = 31.195×e^{(-0.443×(E.V.))}, where the molecular weight (MW) is given in kDa and the elution volume (E.V.) is given in mL; the correlation coefficient for this relationship (R²) is 0.987 (blue line).

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