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Review
. 2022 Oct 31;50(5):1403-1414.
doi: 10.1042/BST20220591.

Mechanistic insights into protein folding by the eukaryotic chaperonin complex CCT

Theresa M Smith  1 , Barry M Willardson  1
Affiliations
Review

Mechanistic insights into protein folding by the eukaryotic chaperonin complex CCT

Theresa M Smith et al. Biochem Soc Trans. .

Abstract

The cytosolic chaperonin CCT is indispensable to eukaryotic life, folding the cytoskeletal proteins actin and tubulin along with an estimated 10% of the remaining proteome. However, it also participates in human diseases such as cancer and viral infections, rendering it valuable as a potential therapeutic target. CCT consists of two stacked rings, each comprised of eight homologous but distinct subunits, that assists the folding of a remarkable substrate clientele that exhibits both broad diversity and specificity. Much of the work in recent years has been aimed at understanding the mechanisms of CCT substrate recognition and folding. These studies have revealed new binding sites and mechanisms by which CCT uses its distinctive subunit arrangement to fold structurally unrelated substrates. Here, we review recent structural insights into CCT-substrate interactions and place them into the broader context of CCT function and its implications for human health.

Keywords: cryo-electron microscopy; molecular chaperones; molecular mechanisms; protein conformation.

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Conflict of interest statement

The authors declare that there are no competing interests associated with the manuscript.

Figures

Figure 1.
Figure 1.. Structure of CCT.
(A) Side and end-on views of human CCT with and without ATP (adapted from PDB 7NVN [32] and 6QB8 [12], respectively), highlighting the double ring arrangement of the eight subunits and the conformational change that occurs in the ATP hydrolysis transition state to cap the central folding chambers of CCT. (B) A ribbon view of the CCT1 subunit showing the domain structure common to all the subunits and the conformational change between the nucleotide-free (gray) and ATP hydrolysis transition state (red) that closes the CCT folding chamber.
Figure 2.
Figure 2.. Functional differences between the CCT hemispheres.
(A) Open form of human CCT (PDB 6QB8) showing the hemispheres centered around CCT2 and CCT6 with high ATPase activity in the CCT2 hemisphere and low ATPase activity in the CCT6 hemisphere. (B) Electrostatic surface views of the interior of the closed form of human CCT (PDB 7NVN) with the CCT6 hemisphere on the left and the CCT2 hemisphere on the right. (PDB 7NVN, positive charge = blue, neutral = white, and negative charge = red.) For the CCT6 hemisphere, the CCT2, 4, 5 and 7 subunits have been removed to reveal the interior of the folding chambers. Positively charged patches in the upper and lower chambers and between the CCT rings are highlighted by yellow ovals. For the CCT2 hemisphere, the CCT1, 3, 6 and 8 subunits have been removed to view the interior of the folding chambers. Negatively charged patches in the upper and lower chambers are highlighted by yellow ovals.
Figure 3.
Figure 3.. Substrate binding positions within CCT.
(A) Side and end-on cut away views of mLST8 (PDB 4JT6) sitting between the rings in the open form of human CCT (PDB 6QB8) [12]. (B) Side and end-on cut away views of β-tubulin in the folding cavity of closed CCT (PDB 7NVN) [32], highlighting the position of tubulin bound in the apical domains of the CCT6 hemisphere. (C) Side and end-on cut away views of actin in the folding cavity of closed CCT (PDB 7NVM) [32], showing actin spanning the cavity to interact with both hemispheres. Substrates are shown in red.
Figure 4.
Figure 4.. Folding trajectory of tubulin.
(A) Model showing the position of tubulin within the open (left) and closed (right) conformations of CCT. In the open form, tubulin is in an amorphous state interacting with the N- and C-termini of the CCT subunits (black lines). In the closed form, the tubulin is released from between the CCT rings as the termini redistribute. The tubulin moves into the CCT folding chamber and interacts with positively charged patches in the apical domains along the inner walls of the chamber. (B) Model representing the four observed states of tubulin folding inside the folding cavity of closed CCT (PDB 7NVN). The domains of tubulin are indicated: N = the N terminal domain, C = the C terminal domain, Core = the helical core domain, Mid = the middle domain. Lines represent unstructured regions in each folding state. See text for details.
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