Supramolecular protein polymers using mini-ferritin Dps as the building block

Area of Science:

• Biochemistry
• Materials Science
• Structural Biology

Background:

• DNA-binding proteins from starved cells (Dps) are mini-ferritins involved in iron homeostasis.
• Engineering Dps proteins can lead to novel biomaterials with enhanced functions.
• The N-terminal region of Dps proteins offers potential sites for modification.

Purpose of the Study:

• To develop a supramolecular assembly using a modified Dps protein from Marinobacter hydrocarbonoclasticus.
• To enhance the iron uptake capability of the Dps protein through supramolecular polymerization.
• To investigate the structural integrity and physiological function of the resulting protein polymers.

Main Methods:

• Site-directed mutagenesis to introduce a cysteine residue (T10C) in the Dps protein N-terminus.
• Click chemistry (thiol-ene coupling) with different linkers to form supramolecular polymers.
• Electrophoresis and mass spectrometry to confirm functionalization and coupling.
• Circular dichroism (CD and SRCD) for secondary structure analysis.
• Dynamic light scattering (DLS), size exclusion chromatography (SEC), and atomic force microscopy (AFM) for size and morphology.
• UV-Vis spectroscopy for assessing iron uptake capacity.

Main Results:

• Successful synthesis of two distinct mini-ferritin supramolecular polymers via thiol-ene click chemistry.
• The DpsT10C mutation and subsequent polymerization did not disrupt the protein's secondary structure.
• The synthesized supramolecular polymers retained the native iron uptake physiological function.
• Characterization confirmed the successful functionalization, coupling, and structural integrity of the polymers.

Conclusions:

• Engineered DpsT10C mini-ferritin can be effectively polymerized into supramolecular structures.
• These novel protein polymers exhibit enhanced iron uptake capabilities.
• The study demonstrates a viable strategy for creating functional protein-based nanomaterials.