PROTAC (Proteolysis-Targeting Chimera), protein hydrolysis targeting chimeras, which use the ubiquitin-protease system to target specific proteins for intracellular degradation, have become an emerging drug modality that has been rapidly developed in recent years.

The PROTAC molecule consists of a target protein ligand (POI ligand), a linker, and an E3 ligand, which specifically binds the target protein while recruiting E3 ligase.

The E3 ligases commonly used in PROTAC are MDM2, cIAP1, VHL and CRBN, and VHL and CRBN are the main ones used in the current study.

CRBN is part of the E3 ubiquitin ligase complex CUL4-RBX1-DDB1, and the common ligand is a dopamine derivative. This derivative has small molecular weight and is good drug-forming, these properties make this protein complex the most studied PROTAC ligase.

We expressed and purified CRBN-DDB1 complex by insect expression system, which can be used for structural study of target - PROTAC molecule - E3 ligase.

The VHL gene is a tumor suppressor gene, and its protein product, VHL, can form a VBC (VHL-ElonginB/C-CUL2) complex with ElonginB, ElonginC, and Cullin2 proteins, which belongs to the E3-ubiquitin protease system and mediates the degradation of various proteins in the human body.

We have developed a system for the expression and purification of the VBC (VHL-ElonginB-ElonginC) complex in E. coli.