The Intrinsically Disordered Cytoplasmic Domain of the T-Cell Receptor Zeta Subunit Does not Form Disordered Dimers

Abstract

Intrinsically disordered proteins play important roles in recognition, signal transduction and molecular sorting. They are generally thought to interact via coupled folding and binding, yielding largely ordered interfaces with their binding partners. In contrast, the intrinsically disordered cytoplasmic domain of the T-cell receptor zeta subunit (ζcyt) was reported to form homodimers with a dissociation constant of ∼10μM in the absence of a disorder-to-order transition. This suggested the existence of highly disordered dimers. We show here using analytical ultracentrifugation that ζcyt is a disordered monomer up to at least millimolar concentrations. Paramagnetic relaxation enhancement demonstrates transient intra- rather than intermolecular interactions. Furthermore, even disulfide crosslinking of ζcyt N-termini, in a configuration reminiscent of T cell receptor clustering, fails to lead to an association of protomers. SEC-MALS confirms the monomeric state of ζcyt but reveals a curious concentration-dependent shift of the elution volume of ζcyt that may previously have been interpreted as dimerization. Our data show that ζcyt does not form a highly disordered protein complex but leave open the question as to whether completely disordered dimers or other oligomers exist in nature.View Large Image | View Hi-Res Image | Download PowerPoint Slide