Cooperative amyloid fibre binding and disassembly by the Hsp70 disaggregase

Abstract

Although amyloid fibres are highly stable protein aggregates, a specific combination of human Hsp70 system chaperones can disassemble them, including fibres formed of α‐synuclein, huntingtin, or Tau. Disaggregation requires the ATPase activity of the constitutively expressed Hsp70 family member, Hsc70, together with the J domain protein DNAJB1 and the nucleotide exchange factor Apg2. Clustering of Hsc70 on the fibrils appears to be necessary for disassembly. Here we use atomic force microscopy to show that segments of in vitro assembled α‐synuclein fibrils are first coated with chaperones and then undergo bursts of rapid, unidirectional disassembly. Cryo‐electron tomography and total internal reflection fluorescence microscopy reveal fibrils with regions of densely bound chaperones, preferentially at one end of the fibre. Sub‐stoichiometric amounts of Apg2 relative to Hsc70 dramatically increase recruitment of Hsc70 to the fibres, creating localised active zones that then undergo rapid disassembly at a rate of ~ 4 subunits per second. The observed unidirectional bursts of Hsc70 loading and unravelling may be explained by differences between the two ends of the polar fibre structure. image The Hsc70 chaperone/disaggregase can disassemble amyloid fibres in a process requiring ATP hydrolysis and clustering of Hsc70 on fibres. Here, Hsc70 is found to be preferentially recruited to ends of amyloid fibres by its J‐domain and nucleotide exchange co‐factors, where it then disassembles the bound fibre segments in rapid, unidirectional bursts. Recycling by the nucleotide exchange factor Apg2 concentrates Hsc70 preferentially at one fibre end Real‐time atomic force microscopy shows disassembly of chaperone‐bound segments in sudden, unidirectional bursts Cryo‐electron tomography shows densely packed arrays of chaperones extending from the fibre surface