E. coli Expression System

Case 1:

1. Expression system optimization: host strains, induction temperature, induction time, inducer concentration, and culture media.

Fig. 1: Optimization of Protein Expression Conditions

2. 3C enzyme was used to remove the His-tag from the target protein. The results of enzyme digestion and protein QC are shown in Fig. 2 and Fig. 3.

Fig. 2: Removal of Target Protein His-tag with 3C Enzyme    Fig. 3: QC Analysis of His-tag Removed Protein

Case 2:

Antibody fragments were expressed in E. coli. Due to the presence of disulfide bonds, incorrect folding during expression led to inclusion bodies. By targeting the periplasmic space for expression, properly folded soluble antibody fragments containing disulfide bonds were obtained.

1. Protein expression and QC results are shown in Fig. 4 and Fig. 5.

Fig. 4: Periplasmic Expression of Antibody Fragment    Fig. 5: Reduced and Non-reduced QC of Antibody Fragment

Lane MW: Protein marker. Lane PP: Periplasmic Space

Case 3:

Many proteins expressed in E. coli form inclusion bodies due to the host's limitations. To ensure the functionality of the target protein, inclusion bodies are refolded into correctly folded soluble proteins for activity assays.

1. Protein expression and QC results are shown in Fig. 6 and Fig. 7.

Fig. 6: Optimization of Target Protein Expression    Fig. 7: Refolding QC of Target Protein