Structure and Interactions of the TPR Domain of Sgt2 with Yeast Chaperones and Ybr137wp

Abstract

Small glutamine-rich tetratricopeptide repeat-containing protein 2 (Sgt2) is a multi-module co-chaperone involved in several protein quality control pathways. The TPR domain of Sgt2 and several other proteins, including SGTA, Hop, and CHIP, is a highly conserved motif known to form transient complexes with molecular chaperones such as Hsp70 and Hsp90. In this work, we present the first high resolution crystal structures of Sgt2_TPR alone and in complex with a C-terminal peptide PTVEEVD from heat shock protein, Ssa1. Using nuclear magnetic resonance spectroscopy and isothermal titration calorimetry, we demonstrate that Sgt2_TPR interacts with peptides corresponding to the C-termini of Ssa1, Hsc82, and Ybr137wp with similar binding modes and affinities.

Substrate relay in an Hsp70‐cochaperone cascade safeguards tail‐anchored membrane protein targeting

“…We hypothesized that another chaperone upstream of Sgt2 is required to fulfill this role. Sgt2 contains a conserved TPR domain that binds multiple heat-shock proteins (Wang et al, 2010;Chartron et al, 2011;Krysztofinska et al, 2017). Among them, Hsp70 has been proposed to deliver TAs to Sgt2 (Chartron et al, 2011).…”

“…Generation of a functional Sgt2ÁTA complex has mostly relied on in vitro translation (IVT) in cell lysates that contain endogenous chaperones (Wang et al, 2010;Rao et al, 2016) or the use of super-physiological concentrations of Sgt2 (Mateja et al, 2015), raising questions as to whether the currently known factors in the GET pathway are sufficient for efficient TA capture and delivery. Intriguingly, Sgt2 contains a conserved tetratricopeptide repeat (TPR) domain that associates with multiple heat-shock proteins including Hsp70, Hsp90, and Hsp104 (Wang et al, 2010;Chartron et al, 2011;Krysztofinska et al, 2017). Heat-shock cognate protein 70 (Hsc70) has been found to associate with TAs translated in mammalian lysate (Abell et al, 2007), and supplementing Hsc70 enhanced TA insertion into the ER in semi-permeabilized cells (Rabu et al, 2008).…”

Structural and mechanistic basis of the EMC-dependent biogenesis of distinct transmembrane clients

“…The helical fold of EMC2 constitutes the central organizer of this platform, established by five or six TPR motifs in human versus yeast, respectively ( Figure 3C). TPR domains are commonly found mediating protein-protein interactions, and are present in numerous well-characterized chaperoneprotein and other interaction networks (Blatch, 1999;Scheufler, 2000;Schlegel, 2007;Assimon, 2015;Krysztofinska, 2017;Graham, 2019). Yeast EMC2 features a more curved helical arrangement with N-and C-terminal domains in closer proximity to each other than seen in human EMC2.…”

The roles of cytosolic quality control proteins, SGTA and the BAG6 complex, in disease

“…Typically TPR domains (Darby et al, 2014); TPR domain (red) residues 84-210 from PDB: 2VYI (Dutta & Tan, 2008). There is currently no solved structure for the C-terminal domain (blue) which encompasses NNP and Q-rich regions (Martinez-Lumbreras et al, 2018). are made up of between 3 and 16 tandem repeats-SGTA contains 3 repeats with each repeat containing 2 almost identically structured antiparallel folded alpha helices (Dutta & Tan, 2008;Krysztofinska et al, 2017). There is additionally a seventh alpha helix known as the C-terminal capping helix that aligns against the second helix of the third repeat, helix 6.…”