PR65, the HEAT-repeat scaffold of phosphatase PP2A, is an elastic connector that links force and catalysis

Abstract

PR65 is the two-layered (α-α solenoid) HEAT-repeat (Huntingtin, elongation factor 3, a subunit of protein phosphatase 2A, PI3 kinase target of rapamycin 1) scaffold of protein phosphatase PP2A. Molecular dynamics simulations predict that, at forces expected in living systems, PR65 undergoes (visco-)elastic deformations in response to pulling/pushing on its ends. At lower forces, smooth global flexural and torsional changes occur via even redistribution of stress along the hydrophobic core of the molecule. At intermediate forces, helix-helix separation along one layer ("fracturing") leads to global relaxation plus loss of contact in the other layer to unstack the affected units. Fracture sites are determined by unusual sequences in contiguous interhelix turns. Normal mode analysis of the heterotrimeric PP2A enzyme reveals that its ambient conformational fluctuations are dominated by elastic deformations of PR65, which introduce a mechanical linkage between the separately bound regulatory and catalytic subunits. PR65-dominated fluctuations of PP2A have the effect of opening and closing the enzyme's substrate binding/catalysis interface, as well as altering the positions of certain catalytic residues. These results suggest that substrate binding/catalysis are sensitive to mechanical force. Force could be imposed from the outside (e.g., in PP2A's response to spindle tension) or arise spontaneously (e.g., in PP2A's interaction with unstructured proteins such as Tau, a microtubule-associated Alzheimer's-implicated protein). The presented example supports the view that conformation and function of protein complexes can be modulated by mechanical energy inputs, as well as by chemical energy inputs from ligand binding. Given that helical-repeat proteins are involved in many cellular processes, the findings also encourage the view that mechanical forces may be of widespread importance.helical-repeat protein | protein elasticity | protein mechanotransduction | spindle tension

Control of mTORC1 signaling by the Opitz syndrome protein MID1

“…Furthermore, an additional distinction of this regulatory loop from the previously described role of PP2A on downstream mTORC1 substrates is that the PP2A holoenzyme responsible for mTORC1 disruption involves the PP2A-Bα (PR55α) subunit rather than the PP2A-B′γ (PR61γ/PP2R5C) subunit implicated in S6K1 dephosphorylation (40). We also found that the association between PP2A-C and mTOR is mediated via the mTOR HEAT repeats, a motif found in Huntingtin, elongation factor 3, A subunit of PP2A, and TOR (41,42). Furthermore, binding of PP2A-C to the activated mTOR mutant L1460P is reduced compared with wild-type mTOR, accounting for the ability of this mutant to rescue mTORC1 signaling in OS cells.…”

ATM and ATR signaling at a glance

“…The N-terminal HEAT repeats act as a scaffold; they mediate the interactions of ATM and ATR with proteins that regulate their catalytic activity and have an important role in their stability (see poster) (Perry and Kleckner, 2003). The HEAT repeats can also act as elastic connectors that undergo deformation following mechanical stimulation and regulate protein activity (Grinthal et al, 2010). As is the case for other PI3K-family members, PIKKs exist as homo-or heterodimers, and dimerization influences their stability and kinase activity.…”

Structural and mechanistic insights into human splicing factor SF3b complex derived using an integrated approach guided by the cryo-EM density maps

“…Large scale changes in curvature are not uncommon for HEAT repeat structures, as has been shown in a number of molecular dynamics studies. 69,70 Thus, it is likely that the partly disordered SF3b155 region (HEAT repeats) and its interactions with all other SF3b proteins (Fig. 1D) constrains the HEAT repeat structure, thereby, forcing it to take such a peculiar curvature.…”

“…S17), this protein has the ability to act as a "molecular sling shot" in the SF3b complex, utilizing its stored elastic potential energy as a wound spring during the transformation of SF3b from closed to the open state. 69,72 Figure 6. Intrinsically disordered region in the SF3b cryo-EM map.…”