Cytosolic chaperonin protects folding intermediates of Gβ from aggregation by recognizing hydrophobic β-strands

Abstract

Cytosolic chaperonin containing t-complex polypeptide 1 (CCT)/TRiC is a group II chaperonin that assists in the folding of newly synthesized proteins. It is a eukaryotic homologue of the bacterial group I chaperonin GroEL. In contrast to the well studied functions of GroEL, the substrate recognition mechanism of CCT/TRiC is poorly understood. Here, we established a system for analyzing CCT/TRiC functions by using a reconstituted protein synthesis by using recombinant elements system and show that CCT/TRiC strongly recognizes WD40 proteins particularly at hydrophobic β-strands. Using the G protein β subunit (Gβ), a WD40 protein that is very rich in β-sheets, as a model substrate, we found that CCT/TRiC prevents aggregation and assists in folding of Gβ, whereas GroEL does not. Gβ has a seven-bladed β-propeller structure; each blade is formed from a WD40 repeat sequence encoding four β-strands. Detailed mutational analysis of Gβ indicated that CCT/TRiC, but not GroEL, preferentially recognizes hydrophobic residues aligned on surfaces of β-strands in the second WD40 repeat of Gβ. These findings indicate that one of the CCT/TRiC-specific targets is hydrophobic β-strands, which are highly prone to aggregation.