It takes a dimer to tango: Oligomeric small heat shock proteins dissociate to capture substrate

Abstract

Small heat-shock proteins (sHsps) are ubiquitous molecular chaperones, and sHsp mutations or altered expression are linked to multiple human disease states. sHsp monomers assemble into large oligomers with dimeric substructure, and the dynamics of sHsp oligomers has led to major questions about the form that captures substrate, a critical aspect of their mechanism of action. We show here that substructural dimers of two plant dodecameric sHsps, Ta16.9 and homologous Ps18.1, are functional units in the initial encounter with unfolding substrate. We introduced inter-polypeptide disulfide bonds at the two dodecameric interfaces, dimeric and nondimeric, to restrict how their assemblies can dissociate. When disulfide-bonded at the nondimeric interface, mutants of Ta16.9 and Ps18.1 (Ta CT-ACD and Ps CT-ACD) were inactive but, when reduced, had WT-like chaperone activity, demonstrating that dissociation at nondimeric interfaces is essential for sHsp activity. Moreover, the size of the Ta CT-ACD and Ps CT-ACD covalent unit defined a new tetrahedral geometry for these sHsps, different from that observed in the Ta16.9 X-ray structure. Importantly, oxidized Ta dimer (disulfide bonded at the dimeric interface) exhibited greatly enhanced ability to protect substrate, indicating that strengthening the dimeric interface increases chaperone efficiency. Temperature-induced size and secondary structure changes revealed that folded sHsp dimers interact with substrate and that dimer stability affects chaperone efficiency. These results yield a model in which sHsp dimers capture substrate before assembly into larger, heterogeneous sHspsubstrate complexes for substrate refolding or degradation, and suggest that tuning the strength of the dimer interface can be used to engineer sHsp chaperone efficiency.

Single-molecule fluorescence-based approach reveals novel mechanistic insights into small heat shock protein chaperone function

“…We demonstrate that initially smaller species of Bc (predominantly monomers and dimers) bind to heat-destabilised CLIC1 oligomers. This observation validates previous suggestions, based on studying end-stage complexes, that smaller species of sHsps have high chaperone ability and therefore initially bind to misfolded proteins 18,42,43 . Interestingly, we observed that the number of complexes formed between Bc and CLIC1 increased rapidly over the first hour of incubation and reached a plateau after 4 hr.…”

“…Whilst it is well established that sHsps can form high-molecular mass complexes with misfolded clients to prevent their aggregation [12][13][14] , little is known about how these complexes are assembled. It has been postulated that smaller sHsp oligomers have enhanced chaperone activity as a result of their increased exposed hydrophobicity and, therefore, a greater affinity for misfolded and aggregation-prone proteins [15][16][17][18] ; however, others have suggested that the larger oligomers are chaperone active [19][20][21] . Thus, it remains unclear precisely how sHsps capture misfolded proteins to form the high-molecular mass sHsp-client complexes observed as a result of their chaperone action.…”

The Mitochondrial Small Heat Shock Protein HSP22 from Pea is a Thermosoluble Chaperone Prone to Co-Precipitate with Unfolding Client Proteins

“…Crystal structures further supported that dimers were the building block of most sHSPs [36]. Dimers were proposed to be the form that captures heat-sensitive unfolding proteins [5,81]. However, heat-induced dissociation of oligomers is not a general feature of sHSPs: for example, plant Class II cytosolic sHSP [40], yeast cytosolic HSP42 [34], sugarcane chloroplast, and mitochondria sHSP [55] remained oligomeric upon heat treatment.…”

N- and C-terminal regions of the small heat shock protein IbpA from Acholeplasma laidlawii competitively govern its oligomerization pattern and chaperone-like activity

“…The oligomerization state of sHSPs can be inuenced by the changes in pH, 9 temperature 10 as well as by post-translational modications, such as phosphorylation 11 that affects their affinity to other proteins and chaperone-like activity. 12 This relationship between changes in oligomerization and chaperone-like activity has been clearly shown for the human HSPB1 and HSPB5 as well as for the yeast HSP42. [13][14][15] While the dynamics of sHSPs oligomerization and oligomers structure were analyzed in detail mainly for archaeal and eukaryotic proteins, the common features of sHSPs seem to be universal in all kingdoms of life (for an extensive review, we refer to ref.…”