Interactions between Small Heat Shock Protein Subunits and Substrate in Small Heat Shock Protein-Substrate Complexes

Kenneth L. Friedrich, Kim C. Giese, Nicole R Buan, Elizabeth Vierling

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Abstract

Small heat shock proteins (sHSPs) are dynamic oligomeric proteins that bind unfolding proteins and protect them from irreversible aggregation. This binding results in the formation of sHSP-substrate complexes from which substrate can later be refolded. Interactions between sHSP and substrate in sHSP-substrate complexes and the mechanism by which substrate is transferred to ATP-dependent chaperones for refolding are poorly defined. We have established C-terminal affinity-tagged sHSPs from a eukaryote (pea HSP18.1) and a prokaryote (Synechocystis HSP16.6) as tools to investigate these issues. We demonstrate that sHSP subunit exchange for HSP18.1 and HSP16.6 is temperature-dependent and rapid at the optimal growth temperature for the organism of origin. Increasing the ratio of sHSP to substrate during substrate denaturation decreased sHSP-substrate complex size, and accordingly, addition of substrate to pre-formed sHSP-substrate complexes increased complex size. However, the size of pre-formed sHSP-substrate complexes could not be reduced by addition of more sHSP, and substrate could not be observed to transfer to added sHSP, although added sHSP subunits continued to exchange with subunits in sHSP-substrate complexes. Thus, although some number of sHSP subunits within complexes remain dynamic and may be important for complex structure/solubility, association of substrate with the sHSP does not appear to be similarly dynamic. These observations are consistent with a model in which ATP-dependent chaperones associate directly with sHSP-bound substrate to initiate refolding.

Original languageEnglish (US)
Pages (from-to)1080-1089
Number of pages10
JournalJournal of Biological Chemistry
Volume279
Issue number2
DOIs
StatePublished - Jan 9 2004

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Small Heat-Shock Proteins
Protein Subunits
Substrates
Adenosine Triphosphate
Synechocystis
Protein Unfolding
Denaturation
Temperature

ASJC Scopus subject areas

  • Biochemistry

Cite this

Interactions between Small Heat Shock Protein Subunits and Substrate in Small Heat Shock Protein-Substrate Complexes. / Friedrich, Kenneth L.; Giese, Kim C.; Buan, Nicole R; Vierling, Elizabeth.

In: Journal of Biological Chemistry, Vol. 279, No. 2, 09.01.2004, p. 1080-1089.

Research output: Contribution to journalArticle

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