Mammalian 20S proteasome isotypes: from structure and function to selective inhibition

• Applicant: Professor Dr. Michael Groll
• Subject Area: Structural Biology
• Term: from 2014 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 256398100

The 20S proteasome core particle (CP) is the key player of the non-lysosomal protein degradation pathway, controls a plethora of essential intracellular processes and is a validated drug target. While simple eukaryotes express only one type of CP, the evolution of mammals and their increased complexity led to diversification and differentiation of the CP, ultimately resulting in four isotypes. Present in all cells, the constitutive proteasome (cCP) accomplishes the bulk of polypeptide degradation required for protein homoeostasis and survival. In contrast, the immunoproteasome (iCP) and the thymoproteasome (tCP) are engaged in shaping a functional adaptive immune system, whereas the role of the spermatoproteasome (sCP) is yet to be elucidated. Although the overall architecture of CP isotypes is identical, their subunit composition varies. Inhibitors that do not discriminate between CP isotypes are blockbuster drugs for the treatment of blood cancers and additional medicinal applications of iCP-selective compounds, e.g. chronic inflammatory or autoimmune diseases, are explored in clinical trials. During the current funding period extensive structure-based inhibitor and mutagenesis studies were performed on cCP and iCP, the results of which we have reported in 11 publications. To date, both tCP and sCP are poorly understood and their structural examination would provide a more thorough understanding of the unique subunits ß5t and Alpha 4s, respectively. Therefore, in the applied follow-up project we aim to structurally and biochemically characterize the tCP. In particular, the analysis of cleavage patterns of tCP versus cCP and iCP will be used to develop tCP-specific ligands. Such compounds may serve future efforts to probe the biological function of the tCP and any medicinal relevance of its selective inactivation. Similar to our work on the iCP, yeast mutagenesis efforts are envisioned to determine crucial elements of ß5t for assembly and function of the tCP. Besides, elaborate experiments on the sCP are planned. The primary goal is expression, purification and crystallization of the genuine human sCP or of a chimeric CP, i.e. a yeast CP that contains the human sCP-specific α4s entity. The envisioned interdisciplinary concept of this proposal certainly will enrich basic proteasome research and may open up future directions in CP drug development. The excellent infrastructure and the long lasting expertise of my lab allow to expect also promising new insights into 20S proteasome isotypes.

• DFG Programme: Research Grants