The vast structural diversity of proteins is responsible for their functional versatility, but also makes their expression and purification a demanding task. Tagging proteins with affinity tags can overcome the complexity of conventional purification by enabling affinity chromatography. An overview will be given for the purification of tagged proteins and then a purification system using a circular permutated caspase will be described in detail. It has been developed for process fusion protein processing to generate an authentic N-terminus for all 20 amino acids. The system employs widely available affinity chromatography resins, and a simple process can be used regardless of the specific nature of the protein of interest.
1. Explain the processing of fusion proteins and non-mAb proteins.
2. Discuss the generation of an authentic N-terminus.
3. Discuss the effect of enzyme kinetics on process of fusion proteins.