Intrinsic Disorder in a Well-Folded Globular Protein

The folded structure of the heterodimeric sweet protein monellin mimics single-chain proteins with topology β1-α1-β2-β3-β4-β5 (chain A: β3-β4-β5; chain B: β1-α1-β2). Furthermore, like naturally occurring single-chain proteins of a similar size, monellin folds cooperatively with no detectable intermediates. However, the two monellin chains, A and B, are marginally structured in isolation and fold only upon binding to each other. Thus, monellin presents a unique opportunity to understand the design of intrinsically disordered proteins that fold upon binding. Here, we study the folding of a single-chain variant of monellin (scMn) using simulations of an all heavy-atom structure-based model. These simulations can explain mechanistic details derived from scMn experiments performed using several different structural probes. scMn folds cooperatively in our structure-based simulations, as is also seen in experiments. We find that structure formation near the transition-state ensemble of scMn is not uniformly distributed but is localized to a hairpin-like structure which contains one strand from each chain (β2, β3). Thus, the sequence and the underlying energetics of heterodimeric monellin promote the early formation of the interchain interface (β2-β3). By studying computational scMn mutants whose "interchain" interactions are deleted, we infer that this energy distribution allows the two protein chains to remain largely disordered when this interface is not folded. From these results, we suggest that cutting the protein backbone of a globular protein between residues which lie within its folding nucleus may be one way to construct two disordered fragments which fold upon binding.