Design of metal-mediated protein assemblies via hydroxamic acid functionalities

Area of Science:

• Biochemistry
• Materials Science
• Synthetic Biology

Background:

• Protein self-assembly is fundamental to life, driving efforts to create synthetic protein architectures.
• Natural protein assemblies rely on complex noncovalent interactions, challenging to replicate in de novo design.
• Developing precise methods for de novo multiprotein system construction is crucial.

Purpose of the Study:

• To present a protocol for metal-mediated protein assembly using hydroxamate (HA) motifs.
• To demonstrate the construction of polyhedral protein cages and 3D crystalline protein-metal-organic frameworks (protein-MOFs).
• To enable graduate-level researchers to synthesize and analyze these novel protein assemblies within six weeks.

Main Methods:

• Utilizing metal-coordinating hydroxamate (HA) motifs for directed protein assembly.
• Employing an asymmetric cytochrome cb562 monomer with Fe3+ and Zn2+ for polyhedral cage formation.
• Synthesizing ditopic HA linkers to create crystalline 3D protein-MOF lattices with protein nodes.

Main Results:

• Successful formation of polyhedral protein cages via concurrent metal ion association.
• Construction of crystalline 3D protein-MOF lattices using bridging HA linkers and protein nodes.
• Characterization of protein cages and protein-MOFs using techniques like X-ray diffraction and electron microscopy.

Conclusions:

• Hydroxamate-mediated assembly provides a robust strategy for constructing sophisticated protein architectures.
• This protocol facilitates the creation of novel protein-based materials with potential functional properties.
• The described methods are accessible for graduate researchers, enabling rapid advancement in protein engineering.