Defining Protein Interactions With Small Heat Shock Proteins

Abstract

Small heat shock proteins (sHSPs) are a class of molecular chaperones broadly observed across organisms where they play a central role in maintaining protein homeostasis under conditions of cellular stress. sHSPs are known to make protein-protein interactions that can be defined into two classes: 1) interactions with unfolded and aggregate-prone proteins (clients) to delay protein aggregation and 2) interactions with each other, leading to the formation of sHSPs oligomers. Here, progress in characterizing details of these interactions is presented. sHSP are defined by three regions: an N-terminal region (NTR), a conserved Alpha Crystallin Domain (ACD), and a C-terminal region (CTR). All three regions are involved in sHSP oligomer formation. Here, we demonstrate the peptides mimicking CTRs of human sHSPs are able to interact with isolated ACDs. Using a strategy based on this observation, we are able to characterize the requirements for ACD/CTR binding and demonstrate this interaction can occur the context of human sHSP heterooligomers. Further, though other interactions are involved in forming sHSP oligomers, we are able to demonstrate ACD/CTR interactions plays a key role in recruiting an incoming subunit to a sHSP oligomer. These observations are relevant to understanding the details dynamic subunit exchange and heterooligomer formation in human sHSPs. Defining how sHSPs interact with clients remains one of the pressing challenges in the sHSP field. It is unclear whether sHSPs interact with clients through a conserved mechanism. Using a variety of techniques, we set to compare and contrast how the human sHSP, HSPB5, and the wheat sHSP, wHSP16.9, interact with clients. Our observations suggest these sHSPs interact with clients through distinct binding modes. Further, the techniques presented were used to characterize other client/sHSP interactions including, client interactions with the HSPB5 disease-associated mutation, R120G.