HSPA1A conformational mutants reveal a conserved structural unit in Hsp70 proteins.
Biochim Biophys Acta Gen Subj
; 1864(1): 129458, 2020 01.
Article
en En
| MEDLINE
| ID: mdl-31676290
ABSTRACT
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, I515, 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. GENERALSIGNIFICANCE:
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.Palabras clave
Texto completo:
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Bases de datos:
MEDLINE
Asunto principal:
Conformación Proteica
/
Adenosina Trifosfato
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Proteínas HSP70 de Choque Térmico
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Regulación Alostérica
Límite:
Humans
Idioma:
En
Revista:
Biochim Biophys Acta Gen Subj
Año:
2020
Tipo del documento:
Article
País de afiliación:
República Checa