SGTA interacts with the proteasomal ubiquitin receptor Rpn13 via a carboxylate clamp mechanism

Abstract

The fate of secretory and membrane proteins that mislocalize to the cytosol is decided by a collaboration between cochaperone SGTA (small, glutamine-rich, tetratricopeptide repeat protein alpha) and the BAG6 complex, whose operation relies on multiple transient and subtly discriminated interactions with diverse binding partners. These include chaperones, membrane-targeting proteins and ubiquitination enzymes. Recently a direct interaction was discovered between SGTA and the proteasome, mediated by the intrinsic proteasomal ubiquitin receptor Rpn13. Here, we structurally and biophysically characterize this binding and identify a region of the Rpn13 C-terminal domain that is necessary and sufficient to facilitate it. We show that the contact occurs through a carboxylate clamp-mediated molecular recognition event with the TPR domain of SGTA, and provide evidence that the interaction can mediate the association of Rpn13 and SGTA in a cellular context.

In silico identification and biochemical characterization of the human dicarboxylate clamp TPR protein interaction network

“…Various ubiquitin C‐terminal hydrolases, E3 ubiquitin–protein ligases, subunits of the proteasome and proteasome interacting proteins as well as ubiquitin receptor and proteins involved in ubiquitin‐mediated proteolysis have been identified. Rpn13, a ubiquitin receptor that interacts with the dcTPR protein SGTA to facilitate degradation of cytosol mislocalized secretory and membrane proteins , is the only reported example of an interaction between dcTPR proteins and members of the ubiquitin–proteasome system via the dicarboxylate clamp mechanism.…”

“…Rpn13, a ubiquitin receptor that interacts with the dcTPR protein SGTA to facilitate degradation of cytosol mislocalized secretory and membrane proteins [9], is the only reported example of an interaction between dcTPR proteins and members of the ubiquitin-proteasome system via the dicarboxylate clamp mechanism.…”

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

“…Hydrophobic substrates bound to the BAG6 complex can be ubiquitinated by the actions of the E3 ligase RNF126 Rodrigo-Brenni, Gutierrez, & Hegde, 2014) and thus targeted for proteasomal degradation. SGTA can interact with the RPN13 subunit of the 19S regulatory particle of the proteasome through its TPR domain (Leznicki et al, 2015;Thapaliya et al, 2016), which has led to the proposal of an SGTA/BAG6 cycle operating at the proteasome (Leznicki & High, 2012). SGTA hands tail-anchored (TA) proteins over Fig.…”

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

“…The co-chaperone, Small glutamine-rich tetratricopeptide repeat (TPR) protein alpha (SGTA), is involved in the decision to target various misfolded and mislocalized proteins into their appropriate pathways, upstream of either insertion to the endoplasmic reticulum (ER) or degradation (Hessa et al, 2011 ; Leznicki and High, 2012 ; Wunderley et al, 2014 ; Casson et al, 2016 ; Shao et al, 2017 ). SGTA interacts with many proteins and forms transient complexes with chaperones, membrane targeting proteins, and members of the ubiquitin/proteasome system (UPS; Rodrigo-Brenni et al, 2014 ; Leznicki et al, 2015 ; Krysztofinska et al, 2016 ; Thapaliya et al, 2016 ).…”

“…TPR domains are well-known for mediating protein-protein interactions (Das et al, 1998 ). The structure of the human SGTA TPR domain was determined previously by X-ray crystallography (Dutta and Tan, 2008 ) and, like the yeast ortholog, has been reported to interact directly with Hsp70/Hsp90 chaperones, the proteasomal subunit Rpn13 and a variety of disease-related proteins (Buchanan et al, 2007 ; Dutta and Tan, 2008 ; Roberts et al, 2015 ; Thapaliya et al, 2016 ). Moreover, the TPR domain structure (including some additional linker residues at the C-terminal) of the Sgt2 homolog from Aspergillus fumigatus was solved by crystallography (Chartron et al, 2011 ).…”