Optimized E. coli expression strain LOBSTR eliminates common contaminants from His-tag purification

Area of Science:

• Biotechnology
• Molecular Biology
• Protein Biochemistry

Background:

• His-tag affinity purification is a common method for recombinant protein isolation from E. coli.
• A significant challenge is the co-purification of histidine-rich E. coli proteins, reducing target protein purity.

Purpose of the Study:

• To engineer a novel E. coli expression strain, LOBSTR (low background strain), to minimize common protein contaminants during His-tag purification.
• To enhance the purity of recombinant proteins obtained through His-tag purification.

Main Methods:

• Genomic modification of the E. coli BL21(DE3) strain to create LOBSTR.
• Engineering reduced affinities of specific E. coli proteins (encoded by arnA and slyD) to nickel and cobalt resins.

Main Results:

• The LOBSTR strain significantly reduces the co-purification of abundant histidine-rich E. coli contaminants.
• Achieved higher purity of target recombinant proteins compared to standard purification methods.
• Demonstrated the utility of LOBSTR for purifying challenging, low-expressing protein targets.

Conclusions:

• LOBSTR provides a robust solution for improving His-tag purification efficiency and target protein purity.
• This engineered strain facilitates the purification of difficult protein targets without additional purification steps, materials, or costs.
• LOBSTR pushes the boundaries of standard His-tag purification techniques.