Figure 2. Structure of AspBHI.

(a) Four AspBHI monomers, A, B, C and D, form a tetramer. Molecules C and D have mobile C-terminal domains (indicated by a circle). (b) AspBHI tetramer, rotated ~90° from the view of panel (a). (c) For comparison, MspJI has an intact tetramer showing in a similar orientation of panel (a). (d) The disordered C-terminal domains of molecules C and D of AspBHI tetramer were located in the void space along the crystallographic 6-fold axis with a diameter of 100 Å. (e) Elution profile of AspBHI on Superdex 200™10/300 GL (GE Healthcare). The column buffer was 20 mM Tris-HCl (pH 7.5), 300 mM NaCl and 1 mM DTT, and 150 ng of AspBHI was loaded onto the column. The inset shows the standardization of the size exclusion column using a Gel Filtration Markers Kit for Protein Molecular Weights (SIGMA-ALDRICH, Cat. No. MWGF1000) at the time AspBHI was profiled using the same buffer. (f) Monomeric AspBHI contains two domains connected by a linker. (g) AspBHI has a discontinuity in strand β8 owing to the insertion of a 3₁₀ helix (right panel), whereas MspJI has a corresponding 20-residue-long curved strand β8 (left panel). Pairwise sequence alignment is shown above the panels. (h) The 3₁₀ helix of molecule A is involved in the dimer interface with the C-terminal helix αL of molecule B. The amino end of the 3₁₀ helix (Ala149 of molecule A) interacts with the carboxyl end of helix αL (Ser368 of molecule B). Arrows indicate helical dipoles.