The Molecular Chaperone DNAJB6 - A Suppressor of Disease Related Amyloid Fibril Formation

Sammanfattning: Several neurodegenerative diseases are caused by peptides or proteins forming amyloid fibrils such as the A? peptide involved in Alzheimer’s disease and the huntingtin exon-1 with a prolonged polyglutamine (polyQ) stretch involved in Huntington’s disease. The ability to form amyloid fibrils is an intrinsic feature in all proteins. The fibrils are built up of monomers to form elongated structures with ?-sheets perpendicular to the fiber axis. Molecular chaperones are involved in maintaining the protein homeostasis in the cells, in preventing protein aggregation, in refolding misfolded proteins and in mediating degradation of proteins and also in preventing amyloid fibril formation. DNAJB6 is a member of the HSP40 chaperone family, and has been found to be superior compared to other chaperones in its ability to prevent aggregation of huntingtin exon-1 in human cells. In this thesis we have expressed and purified DNAJB6 in order to characterize its structure and function. Our data show that DNAJB6 also in vitro can suppress the fibril formation of polyQ and A? peptides by a direct interaction. By kinetic analysis of the nucleation rate constants we conclude that there is a strong binding between DNAJB6 and the early aggregated species. This prevents them from growing further and thereby efficiently inhibiting fibril formation. It was previously suggested that the inhibitory effect was due to a serine and threonine rich region in DNAJB6. We constructed a series of serine and threonine to alanine substitutions mutants of DNAJB6. The potency of DNAJB6 to suppress amyloid fibril formation declined in proportion to the number of substitutions. Furthermore, a structure model of DNAJB6 was generated and evaluated with chemical crosslinking mass spectrometry. The model revealed a possible domain arrangement where the conserved serine- and threonine-rich region forms a disordered protruding domain located between the well-ordered N- and C-terminal domains. The results obtained in this thesis suggest that DNAJB6 is an efficient suppressor of amyloid fibril formation, acting early in the self-assembly process. Since DNAJB6 is expressed in the brain and can prolong the life-span of a Huntington’s disease mouse model, it is a potential therapeutic target for neurodegenerative diseases.