Structural resemblance of the DNAJA-family protein, Tid1, to the DNAJB-family Hsp40.

The specific pair of heat shock protein 70 (Hsp70) and Hsp40 constitutes an essential molecular chaperone system involved in numerous cellular processes, including the proper folding/refolding and transport of proteins. Hsp40 family members are characterized by the presence of a conserved J-domain (JD) that functions as a co-chaperone of Hsp70. Tumorous imaginal disc 1 (Tid1) is a tumor suppressor protein belonging to the DNAJA3 subfamily of Hsp40 and functions as a co-chaperone of the mitochondrial Hsp70, mortalin. In this work, we performed nuclear magnetic resonance spectroscopy to determine the solution structure of JD and its interaction with the glycine/phenylalaninerich region (GF-motif) of human Tid1. Notably, Tid1-JD, whose conformation was consistent with that of the DNAJB1 JD, appeared to stably interact with its subsequent GF-motif region. Collectively with our sequence analysis, the present results demonstrate that the functional and regulatory mode of Tid1 resembles that of the DNAJB1 subfamily members rather than DNAJA1 or DNAJA2 subfamily proteins. Therefore, it is suggested that an allosteric interaction between mortalin and Tid1 is involved in the mitochondrial Hsp70/Hsp40 chaperone system. [BMB Reports 2022; 55(10): 488-493].

Unique Unfoldase/Aggregase Activity of a Molecular Chaperone Hsp33 in its Holding-Inactive State.

The various chaperone activities of heat shock proteins contribute to ensuring cellular proteostasis. Here, we demonstrate the non-canonical unfoldase activity as an inherent functionality of the prokaryotic molecular chaperone, Hsp33. Hsp33 was originally identified as a holding chaperone that is post-translationally activated by oxidation. However, in this study, we verified that the holding-inactive reduced form of Hsp33 (RHsp33) strongly bound to the translational elongation factor, EF-Tu. This interaction was critically mediated by the redox-switch domain of RHsp33 and the guanine nucleotide-binding domain of EF-Tu. The bound RHsp33, without undergoing any conformational change, catalyzed the EF-Tu aggregation by evoking the aberrant folding of EF-Tu to expose hydrophobic surfaces. Consequently, the oligomers/aggregates of EF-Tu, but not its functional monomeric form, were highly susceptible to proteolytic degradation by Lon protease. These findings present a unique example of an ATP-independent molecular chaperone with distinctive dual functions-as an unfoldase/aggregase and as a holding chaperone-depending on the redox status. It is also suggested that the unusual unfoldase/aggregase activity of RHsp33 can contribute to cellular proteostasis by dysregulating EF-Tu under heat-stressed conditions.

C-terminal dimerization of apo-cyclic AMP receptor protein validated in solution.

Although cyclic AMP receptor protein (CRP) has long served as a typical example of effector-mediated protein allostery, mechanistic details into its regulation have been controversial due to discrepancy between the known crystal structure and NMR structure of apo-CRP. Here, we report that the recombinant protein corresponding to its C-terminal DNA-binding domain (CDD) forms a dimer. This result, together with structural information obtained in the present NMR study, is consistent with the previous crystal structure and validates its relevance also in solution. Therefore, our findings suggest that dissociation of the CDD may be critically involved in cAMP-induced allosteric activation of CRP.

Dimerization of TRAF-interacting protein (TRAIP) regulates the mitotic progression.

The homo- or hetero-dimerization of proteins plays critical roles in the mitotic progression. The TRAF-interacting protein (TRAIP) is crucial in early mitotic progression and chromosome alignment defects in the metaphase. The TRAIP is a 469 amino acid protein, including the Really Interesting New Gene (RING), coiled-coil (CC), and leucine zipper (LZ) domain. In general, the CC or LZ domain containing proteins forms homo- or hetero-dimerization to achieve its activity. In this study, a number of TRAIP mutants were used to define the TRAIP molecular domains responsible for its homo-dimerization. A co-immunoprecipitation assay indicated that the TRAIP forms homo-dimerization through the CC domain. The cells, expressing the CC domain-deleted mutant that could not form a homo-dimer, increased the mitotic index and promoted mitotic progression.