Efficient Purification of Polyhistidine-Tagged Recombinant Proteins Using Functionalized Corundum Particles

Area of Science:

• Biochemistry
• Protein Purification
• Materials Science

Background:

• Immobilized metal affinity chromatography (IMAC) is widely used for polyhistidine-tagged protein purification.
• Traditional IMAC methods face limitations requiring optimization and additional steps.
• A need exists for more efficient, cost-effective, and robust protein purification techniques.

Purpose of the Study:

• To develop and evaluate functionalized corundum particles for efficient recombinant protein purification.
• To assess the performance of corundum-based purification in complex biological matrices and large sample volumes.
• To compare the novel corundum method with standard Ni-NTA agarose beads.

Main Methods:

• Corundum particles were functionalized with APTES, EDTA dianhydride, and loaded with nickel ions.
• Kaiser test and ICP-MS were used to monitor functionalization and quantify metal-binding capacity.
• Purification efficiency was tested using His-tagged protein A/G, E. coli cytoplasm, His6-MBP-mSA2, and SARS-CoV-2-S-RBD-His8.

Main Results:

• Corundum particles demonstrated a binding capacity of approximately 3 mg protein/g.
• Higher purity was achieved compared to Ni-NTA agarose beads, even with large sample volumes (1 L).
• Cost-effective purification (less than 30 cents/g) with high physical and chemical stability was achieved.

Conclusions:

• Functionalized corundum particles provide an efficient, robust, and cost-effective platform for His-tagged protein purification.
• The column-free method is suitable for complex matrices, large volumes, and low protein concentrations.
• This novel material has potential for both laboratory and industrial-scale applications.