Intrinsically disordered proteins (IDPs) studied by EPR and in-cell EPR

Sabrina Weickert, Julia Cattani and Malte Drescher

DOI: 10.1039/9781788013888-00001

Intrinsically disordered proteins (IDPs) play important physiological, but also disease-related roles. In order to understand the function and malfunction of proteins of this class, electron paramagnetic resonance (EPR) spectroscopy has proven to be a valuable tool, allowing investigation of the protein structural ensembles upon interaction with the environment. This review focuses on the IDPs tau and a-synuclein and gives an overview over recent EPR studies performed with these proteins.

1 Introduction

1.1 IDPs

Intrinsically disordered proteins (IDPs) are a class of proteins lacking a stable three-dimensional structure in solution. Nonetheless, these proteins are native and fulfill many important biological functions, among them cell signaling, recognition and regulation. IDPs are highly prevalent in humans: Genome analyses predict that around 25% of all human proteins are disordered from end to end, while up to 40% contain unstructured regions. Due to their abundance as well as their unique structural and dynamical flexibility, IDPs are key players in many biological pathways, capable of specific interactions with a multitude of binding partners and therefore often binding to or serving as hubs in protein interaction networks. Due to the same reasons, many members of the IDP family are known to be associated with a variety of human diseases, among them prominently cancers, cardiovascular diseases, diabetes and neurodegenerative diseases. All of this has made IDPs a research field of tremendous importance and interest since the turn of the century.

1.1.1 The peculiar free energy landscape of IDPs. In the free energy landscape of proteins with a native globular fold there is a pronounced free energy minimum that stabilizes a distinct 3D fold, which often represents the unique functional form of a globular protein (Fig. 1A). The decrease in entropy associated with restrictions of the conformational freedom during folding is compensated by the formation of many intramolecular contacts. However, the free energy landscape of an IDP looks distinctly different (Fig. 1B): It is characterized by the lack of a global free energy minimum, but shows many local minima instead, which are separated by small energy barriers that allow quick and frequent interconversion between the accessible states. Consequently, the conformational ensemble of an IDP in solution is heterogeneous and characterized by a dynamic exchange between many accessible structures. This non-folding behavior is encoded in the amino acid composition of IDPs: These proteins contain less hydrophobic (Ile, Leu, Val) and aromatic (Trp, Tyr, Phe) amino acid residues, but a significantly larger proportion of small and hydrophilic amino acid residues (Arg, Gly, Gln, Ser, Pro, Glu, Lys) and are richer in structure-breaking amino acid residues (Pro, Gly) than typical globular proteins. Although IDPs cannot spontaneously fold into a compact globular structure, the presence of interaction partners can alter the IDP free energy landscape in a way that more pronounced energy minima appear: Upon interaction with a partner, IDPs often undergo a structural reorganization that defines a state of clearly reduced free energy, which is thermodynamically stabilized (Fig. 1B).

1.1.2 Linking folding and binding. While some IDPs undergo a folding process as a whole upon interaction with a binding partner, more commonly specific recognition motifs in the disordered protein adopt secondary or tertiary structural elements upon interaction with a binding partner. As a mechanism of these disorder-to-order transitions, two major models are discussed in the literature – the 'conformational selection' model and the 'induced folding' model. The first model is based on the assumption that the binding partner selects a specific conformation resembling the bound conformation from the wide ensemble of coexisting conformations the IDP adopts when free in solution (Fig. 1B). The latter model is based on the assumption that the IDP binds to its interaction partner in the fully disordered state and folds while bound to the partner, i.e., folding is induced by the partner. In reality, one or the other process may occur, or also some combination of the two models. The full dimension of the structural flexibility of IDPs becomes obvious in cases, where one recognition domain of an IDP can adopt various structural folds upon interaction with different binding partners. One prominent example is the C-terminal disordered region of tumor suppressor p53, which can adopt helical, ß-strand or irregular structure upon interaction with different partners. In contrast to a concise disorder-to-order transition, some IDPs may also stay largely disordered and still show fast interconversion between coexisting conformations while in functional complex with a binding partner. Such heterogeneous protein complexes are referred to as 'fuzzy complexes'.

1.1.3 IDPs in diseases. Similarly to the formation of functional complexes with protein partners, a profound indentation in the free energy landscape of IDPs can be caused by formation of non-functional complexes of IDPs, like oligomers, amorphous...