Studies on the effect of the J-domain on the substrate binding domain (SBD) of Hsp70 using a chimeric human J-SBD polypeptide.

Autor: Tiroli-Cepeda AO; Universidade Estadual de Campinas, Campinas, SP 13083-970, Brazil., Seraphim TV; Universidade Estadual de Campinas, Campinas, SP 13083-970, Brazil., Pinheiro GMS; Universidade Estadual de Campinas, Campinas, SP 13083-970, Brazil., Souto DEP; Universidade Estadual de Campinas, Campinas, SP 13083-970, Brazil., Kubota LT; Universidade Estadual de Campinas, Campinas, SP 13083-970, Brazil; INCTBioanalitica, Brazil., Borges JC; São Carlos Institute of Chemistry, University of São Paulo, São Carlos, SP 13560-970, Brazil., Barbosa LRS; Institute of Physics, University of Sao Paulo USP, São Paulo, SP, Brazil., Ramos CHI; Universidade Estadual de Campinas, Campinas, SP 13083-970, Brazil; INBEB, Brazil. Electronic address: cramos@iqm.unicamp.br.
Jazyk: angličtina
Zdroj: International journal of biological macromolecules [Int J Biol Macromol] 2019 Mar 01; Vol. 124, pp. 111-120. Date of Electronic Publication: 2018 Nov 17.
DOI: 10.1016/j.ijbiomac.2018.11.130
Abstrakt: DnaJ/Hsp40 chaperones deliver unfolded proteins and stimulate the ATPase activity of DnaK/Hsp70 via their J-domain. However, the interaction is transient, creating a challenge for detailed analysis. We investigated whether it would be possible to gain further understanding of this interaction by engineering a chimeric polypeptide where the J-domain of Hsp40 was covalently attached to the substrate binding domain (SBD) of Hsp70 by a flexible linker. The rationale is to increase the proximity between the interacting partners to promote their natural interaction and facilitate the characterization of the interaction. The resulting chimera, termed J-SBD, was properly folded and had properties not present in the full-length Hsp70 or in the SBD alone, for instance a higher protective effect against aggregation and being a monomer. Substrate binding also appear to exceed that of SBD alone as revealed by a decreased binding to bis-ANS, a probe for hydrophobic patches. This hypothesis is supported by the structural model created by small angle X-ray scattering, suggesting that the lid subdomain (SBDα) is partially opened in the J-SBD. Collectively, our results suggest a model in which J-domain binding may shift the Hsp70 equilibrium towards the monomer state, exposing hydrophobic sites prone to substrate accommodation.
(Copyright © 2018 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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