Structural metamorphism and polymorphism in proteins on the brink of thermodynamic stability

Abstract

The classical view of the structure-function paradigm advanced by Anfinsen in the 1960s is that a protein's function is inextricably linked to its three-dimensional structure and is encrypted in its amino acid sequence. However, it is now known that a significant fraction of the proteome consists of intrinsically disordered proteins (IDPs). These proteins populate a polymorphic ensemble of conformations rather than a unique structure but are still capable of performing biological functions. At the boundary, between well-ordered and inherently disordered states are proteins that are on the brink of stability, either weakly stable ordered systems or disordered but on the verge of being stable. In such marginal states, even relatively minor changes can significantly alter the energy landscape, leading to large-scale conformational remodeling. Some proteins on the edge of stability are metamorphic, with the capacity to switch from one fold topology to another in response to an environmental trigger (e.g., pH, temperature/salt, redox). Many IDPs, on the other hand, are marginally unstable such that small perturbations (e.g., phosphorylation, ligands) tip the balance over to a range of ordered, partially ordered, or even more disordered states. In general, the structural transitions described by metamorphic fold switches and polymorphic IDPs possess a number of common features including low or diminished stability, large-scale conformational changes, critical disordered regions, latent or attenuated binding sites, and expansion of function. We suggest that these transitions are, therefore, conceptually and mechanistically analogous, representing adjacent regions in the continuum of order/disorder transitions.

Keywords: fold switching; intrinsically disordered proteins; metamorphic proteins; protein malleability.

Figure 1

Proteins on the brink of stability can undergo a continuum of order/disorder transitions. (A) Examples of transitions from top left to bottom right: Transition between the extended and collapsed disordered states of prostate associated Gene 4 (PAGE4), modulated by phosphorylation;111 disorder‐to‐order transition of 4E‐BP2 induced by phosphorylation;23 order‐to‐order fold switching between G_A98 and G_B98, triggered by single amino acid changes or ligand binding.64 In contrast, stable proteins such as subtilisin (shown in dark blue) do not undergo such changes. (B) Approximate energy well diagrams for each protein from PAGE4 (top) to subtilisin (bottom).

Figure 2

Examples of metamorphic proteins where disordered or partially disordered regions play an important role in remodeling ordered states. N‐ and C‐terminal regions of fold‐switched domains are color coded cyan and red, respectively. All cases are naturally occurring with the exception of (B), which is a designed system. Structures for each panel are identified left to right. (A) Lymphotactin‐10 (PDB 1J8I), Lymphotactin‐40 (PDB 2JP1). (B) GA98 (PDB 2LHC), GB98 (PDB 2LHD). (C) Chloride intracellular channel 1 (CLIC1)‐oxidized (PDB 1RK4), CLIC1‐reduced (PDB 1K0M). (D) P1 lysozyme‐inactive (PDB 1XJU), P1 lysozyme‐active (PDB 1XJT). (E) hemagglutinin pre‐fusion (PDB 5HMG), post‐fusion (PDB 1HTM). (F) T7 RNA polymerase‐initiation state (PDB 1QLN), elongation state (PDB 1MSW).

Figure 3

Latent and attenuated binding sites become accessible in a range of order–order, order–disorder, and disorder–disorder transitions involving metamorphic and polymorphic proteins. (A) The metamorphic protein O‐Mad2 (PDB 1DUJ) has a buried binding site (red) that is only accessible to cdc20 (green) upon fold switching to C‐Mad2 (1S2H). (B) Partially disordered 4E‐BP2 binds to eIF4E (green) utilizing an exposed helical motif (red) that is masked in a β‐sheet structure upon phosphorylation. (C) The disordered protein Myc (1‐88) has multiple transient long‐range interactions that attenuate the affinity of its binding epitope (red) for Bin1‐SH3 (green).

Figure 4

Ensemble switching between relatively closed and open disordered states. (A) Differential phosphorylation remodels the PAGE4 ensemble. Cartoon depiction of the HIPK1‐PAGE4 polypeptide chain (top) showing competing long‐range electrostatic interactions that decrease the radius of gyration of the polypeptide chain. The purple rectangle represents a transient helix. Hyper‐phosphorylation by CLK2 (bottom) weakens these long‐range interactions and decreases the helical propensity, leading to a more extended conformation with larger radius of gyration. (B) Conformational ensembles for HIPK1‐PAGE4 (top) and CLK2‐PAGE4 (bottom) from MD simulations.112