The interaction of the mitochondrial protein importer TOMM34 with HSP70 is regulated by TOMM34 phosphorylation and binding to 14-3-3 adaptors.
J Biol Chem
; 295(27): 8928-8944, 2020 07 03.
Article
em En
| MEDLINE
| ID: mdl-32371396
ABSTRACT
Translocase of outer mitochondrial membrane 34 (TOMM34) orchestrates heat shock protein 70 (HSP70)/HSP90-mediated transport of mitochondrial precursor proteins. Here, using in vitro phosphorylation and refolding assays, analytical size-exclusion chromatography, and hydrogen/deuterium exchange MS, we found that TOMM34 associates with 14-3-3 proteins after its phosphorylation by protein kinase A (PKA). PKA preferentially targeted two serine residues in TOMM34 Ser93 and Ser160, located in the tetratricopeptide repeat 1 (TPR1) domain and the interdomain linker, respectively. Both of these residues were necessary for efficient 14-3-3 protein binding. We determined that phosphorylation-induced structural changes in TOMM34 are further augmented by binding to 14-3-3, leading to destabilization of TOMM34's secondary structure. We also observed that this interaction with 14-3-3 occludes the TOMM34 interaction interface with ATP-bound HSP70 dimers, which leaves them intact and thereby eliminates an inhibitory effect of TOMM34 on HSP70-mediated refolding in vitro In contrast, we noted that TOMM34 in complex with 14-3-3 could bind HSP90. Both TOMM34 and 14-3-3 participated in cytosolic precursor protein transport mediated by the coordinated activities of HSP70 and HSP90. Our results provide important insights into how PKA-mediated phosphorylation and 14-3-3 binding regulate the availability of TOMM34 for its interaction with HSP70.
Palavras-chave
14-3-3 protein; 70-kDa heat shock protein (Hsp70); HSP70; Hsp70; Tomm34; dimerization; hydrogen-deuterium exchange; molecular chaperone; phosphorylation; protein folding; protein import; protein kinase A (PKA); protein-nucleic acid interaction; proteinprotein interaction; translocase of outer mitochondrial membrane 34 (TOMM34)
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Proteínas de Choque Térmico HSP70
/
Proteínas de Transporte da Membrana Mitocondrial
/
Proteínas 14-3-3
Limite:
Humans
Idioma:
En
Revista:
J Biol Chem
Ano de publicação:
2020
Tipo de documento:
Article
País de afiliação:
República Tcheca