A low-cost affinity purification system using β-1,3-glucan recognition protein and curdlan beads

Area of Science:

• Biochemistry
• Molecular Biology
• Protein Engineering

Background:

• Silkworm β-1,3-glucan recognition protein (βGRP) exhibits specific binding to β-1,3-glucan.
• Existing affinity purification systems face limitations in cost and chemical stability.

Purpose of the Study:

• To develop a novel, cost-effective affinity purification system using βGRP.
• To evaluate the efficiency and stability of the new system compared to conventional methods.

Main Methods:

• Development of the GRP system utilizing the βGRP recognition domain (GRP-tag) and curdlan beads.
• Preparation of curdlan beads via alkaline solubilization, neutralization, sonication, and centrifugation.
• Application of the GRP system for recombinant protein purification and comparison with GST-tag system.

Main Results:

• The GRP system demonstrated 2-3 times higher expression levels of GRP-tagged proteins compared to GST-tagged proteins.
• Protein purity achieved with the GRP system was comparable to the GST system.
• The GRP system exhibited superior chemical stability, particularly at low pH (4-6).
• Proteins purified using the GRP system were structurally and functionally active.

Conclusions:

• The GRP system provides an economical and robust method for recombinant protein purification.
• This system is suitable for both large- and small-scale protein preparation for functional and structural studies.
• The GRP system offers advantages in expression levels, stability, and cost-effectiveness over traditional methods.