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

Autor(es)

Tiroli-Cepeda, Ana O. ^[1] ; Seraphim, Thiago V. ^[1] ; Pinheiro, Glaucia M. S. ^[1] ; Souto, Denio E. P. ^[1] ; Kubota, Lauro T. ^{[1, 2]} ; Borges, Julio C. ^[3] ; Barbosa, Leandro R. S. ^[4] ; Ramos, Carlos H. I. ^{[1, 5]}

Afiliação do(s) autor(es)

^[1] Univ Estadual Campinas, BR-13083970 Campinas, SP - Brazil

^[2] INCTBioanalitica, Campinas, SP - Brazil

^[3] Univ Sao Paulo, Sao Carlos Inst Chem, BR-13560970 Sao Carlos, SP - Brazil

^[4] Univ Sao Paulo, Inst Phys, Sao Paulo, SP - Brazil

^[5] INBEB, Londrina - Brazil

• Tipo de documento: Artigo Científico
• Fonte: International Journal of Biological Macromolecules; v. 124, p. 111-120, MAR 1 2019.
• Citações Web of Science: 0

Resumo

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 (SBD alpha) 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. (C) 2018 Elsevier B.V. All rights reserved. (AU)

• Processo FAPESP: 14/07206-6 - Estudos da HSP70 mitocondrial de humanos e de protozoários: abordagem estrutural e funcional
• Beneficiário: Julio Cesar Borges
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 17/07335-9 - Estudos das isoformas da HSP70 humana residentes no citoplasma e mitocôndria e de seus oligômeros de alta massa molecular: interação com co-chaperonas e proteínas clientes
• Beneficiário: Julio Cesar Borges
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 15/15822-1 - Estudo das propriedades físico-químicas e estruturais de fármacos e líquidos iônicos com sistemas de relevância biológica
• Beneficiário: Leandro Ramos Souza Barbosa
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 12/50161-8 - Estudo da estrutura e função da chaperona Hsp90 com ênfase no seu papel em homeostase celular
• Beneficiário: Carlos Henrique Inacio Ramos
• Modalidade de apoio: Auxílio à Pesquisa - Temático