The symmetric designer protein Pizza as a scaffold for metal coordination

Abstract

Symmetric proteins are currently of interest as they allow creation of larger assemblies and facilitate the incorporation of metal ions in the larger complexes. Recently this was demonstrated by the biomineralization of the cadmium-chloride nanocrystal via the Pizza designer protein. However, the mechanism behind this formation remained unclear. Here, we set out to investigate the mechanism driving the formation of this nanocrystal via truncation, mutation, and circular permutations. In addition, the interaction of other biologically relevant metal ions with these symmetric proteins to form larger symmetric complexes was also studied.The formation of the initial nanocrystal is shown to originate from steric strain, where His 58 induces a different rotameric conformation on His 73, thereby distorting an otherwise perfect planar ring of alternating cadmium and chlorine ions, resulting in the smallest nanocrystal.Similar highly symmetric complexes were also observed for the other biological relevant metal ions. However, the flexibility of the coordinating histidine residues allows each metal ion to adopt its preferred geometry leading to either monomeric or dimeric 𝛽𝛽-propeller units, where the metal ions are located at the interface between both propeller units. These results demonstrate that symmetric proteins are not only interesting to generate larger assemblies, but are also the perfect scaffold to create more complex metal based assemblies. Such metal protein assemblies may then find applications in bionanotechnology or biocatalysis.

Hierarchical Self-Assembly of a Supramolecular Protein-Metal Cage Encapsulating a Polyoxometalate Guest

“…The size distribution shifts to an average value of 8 nm, which is consistent with the value previously reported for a Pizza6 dimer . This supports the previously stated hypothesis that an increase in pH above the p K a of the His residues on the surface of Pizza6-S is crucial for the observed complex formation . It should be noted that the resulting complex depends on the synergy of Cu II and the PW9 POM anion since such a size is only observed when both are present via the addition of Cu3 .…”

“…4 This supports the previously stated hypothesis that an increase in pH above the pK a of the His residues on the surface of Pizza6-S is crucial for the observed complex formation. 24 It should be noted that the resulting complex depends on the synergy of Cu II and the PW9 POM anion since such a size is only observed when both are present via the addition of Cu3. A more accurate analysis of the size of this complex via mass spectrometry, e.g., ESI-MS, would be desirable; however, it is not possible due to the presence of the polyoxometalate, which due to its strong charge and molecular weight is incompatible with protein mass spectrometry.…”

“…The protein-POM crystal is solved in the P 2 1 2 1 2 space group at a resolution of 1.80 Å. The single-crystal structure displays a dimeric assembly of Pizza6 proteins, which does not form in the absence of Cu3 . ,, The resulting crystal structure bears the unmistakable presence of POM molecules bound to the protein. However, the assembly that was observed deviates from the expected Cu3 –Pizza6-S complex.…”