Structural basis for the structural dynamics of human mitochondrial chaperonin mHsp60

Abstract

Human mitochondrial chaperonin mHsp60 is essential for mitochondrial function by assisting folding of mitochondrial proteins. Unlike the double-ring bacterial GroEL, mHsp60 exists as a heptameric ring that is unstable and dissociates to subunits. The structural dynamics has been implicated for a unique mechanism of mHsp60. We purified active heptameric mHsp60, and determined a cryo-EM structure of mHsp60 heptamer at 3.4 Å. Of the three domains, the equatorial domains contribute most to the inter-subunit interactions, which include a four-stranded β sheet. Our structural comparison with GroEL shows that mHsp60 contains several unique sequences that directly decrease the sidechain interactions around the β sheet and indirectly shorten β strands by disengaging the backbones of the flanking residues from hydrogen bonding in the β strand conformation. The decreased inter-subunit interactions result in a small inter-subunit interface in mHsp60 compared to GroEL, providing a structural basis for the dynamics of mHsp60 subunit association. Importantly, the unique sequences are conserved among higher eukaryotic mitochondrial chaperonins, suggesting the importance of structural dynamics for eukaryotic chaperonins. Our structural comparison with the single-ring mHsp60-mHsp10 shows that upon mHsp10 binding the shortened inter-subunit β sheet is restored and the overall inter-subunit interface of mHsp60 increases drastically. Our structural basis for the mHsp10 induced stabilization of mHsp60 subunit interaction is consistent with the literature that mHsp10 stabilizes mHsp60 quaternary structure. Together, our studies provide structural bases for structural dynamics of the mHsp60 heptamer and for the stabilizing effect of mHsp10 on mHsp60 subunit association.

Hereditary spastic paraplegia SPG13 mutation increases structural stability and ATPase activity of human mitochondrial chaperonin

“…In the presence of their own cochaperonins, chaperonins display decreased ATPase activities. For example, mHsp60 displays 50% decrease in the ATPase activity in the presence of mHsp10 ( 23 , 26 – 28 , also Fig. 2 C, Table 2 ).…”

“…Unlike the paradigm bacterial chaperonin GroEL, mHsp60 is intrinsically dynamic as evident by its irreversible dissociation to monomers 24 , 25 . In our recent structural analysis 23 , we found that mHsp60 contains unique sequences that weaken the inter-subunit interaction, accounting for dynamics of the subunit association. In this study, we found that the V72I mutation fortuitously fosters inter-subunit interactions including a potential charge-charge interaction between K74 and S44, stabilizing the subunit association.…”

“…2 A, Table 1 ). To confirm the increased stability effect, we removed the last 26 a.a. fragment mainly consisting of the Gly-Met repeats, which is not visible in the CryoEM structure of mHsp60 23 , to generate a C-terminal truncation mHsp60 V72I/GMless . Consistently, mHsp60 V72I/GMless had a higher T m and larger ∆H unfolding than mHsp60 (Fig.…”

“…The equatorial domain contains the ATP binding site, and forms most of the inter-subunit interaction. Our recent structural analysis shows that the inter-subunit interactions in mHsp60 are weaker than those in the bacterial homolog GroEL 23 , providing structural basis for the intrinsic dissociation of the mHsp60 heptamer 24 , 25 . Notably, the SPG13 HSP associated residue V72 is located within the equatorial domain: it is in the middle of a long helix, the 22-a.a. α3 (Fig.…”

“…mHsp60 is homo-heptameric with the seven subunits forming a ring-like structure (Fig. 1 A) 23 . Each subunit consists of three domains, the apical, intermediate and equatorial domains (Fig.…”