HSPA1A conformational mutants reveal a conserved structural unit in Hsp70 proteins
| Autoři | |
|---|---|
| Rok publikování | 2020 |
| Druh | Článek v odborném periodiku |
| Časopis / Zdroj | Biochimica et Biophysica Acta - General Subjects |
| Fakulta / Pracoviště MU | |
| Citace | |
| www | https://doi.org/10.1016/j.bbagen.2019.129458 |
| Doi | http://dx.doi.org/10.1016/j.bbagen.2019.129458 |
| Klíčová slova | Allostery; Molecular chaperones; Heat-shock protein 70; Folding; Mutation |
| Popis | Background: The Hsp70 proteins maintain proteome integrity through the capacity of their nucleotide- and substrate-binding domains (NBD and SBD) to allosterically regulate substrate affinity in a nucleotide-dependent manner. Crystallographic studies showed that Hsp70 allostery relies on formation of contacts between ATP-bound NBD and an interdomain linker, accompanied by SBD subdomains docking onto distinct sites of the NBD leading to substrate release. However, the mechanics of ATP-induced SBD subdomains detachment is largely unknown. Methods: Here, we investigated the structural and allosteric properties of human HSPA1A using hydrogen/deuterium exchange mass spectrometry, ATPase assays, surface plasmon resonance and fluorescence polarization-based substrate binding assays. Results: Analysis of HSPA1A proteins bearing mutations at the interface of SBD subdomains close to the interdomain linker (amino acids L399, L510, 1515, and D529) revealed that this region forms a folding unit stabilizing the structure of both SBD subdomains in the nucleotide-free state. The introduced mutations modulate HSPA1A allostery as they localize to the NBD-SBD interfaces in the ATP-bound protein. Conclusions: These findings show that residues forming the hydrophobic structural unit stabilizing the SBD structure are relocated during ATP-activated detachment of the SBD subdomains to different NBD-SBD docking interfaces enabling HSPA1A allostery. General significance: Mutation-induced perturbations tuned HSPA1A sensitivity to peptide/protein substrates and to Hsp40 in a way that is common for other Hsp70 proteins. Our results provide an insight into structural rearrangements in the SBD of Hsp70 proteins and highlight HSPA1A-specific allostery features, which is a prerequisite for selective targeting in Hsp-related pathologies. |
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