Biophysical characterization of a recombinant soluble interleukin 2 receptor (Tac). Evidence for a monomeric structure.

The interleukin 2 receptor (IL2R) plays a prominent role in the biology of T cells, B cells, and NK cells during activation. Of the three chains described, the alpha-chain of the receptor (Tac; IL2R alpha; CD25) is the most subject to regulation and is shed from the surface of activated cells to generate a soluble form in serum and tissues. Conflicting results have been reported on the native structure of soluble Tac, suggesting variously a monomer, a dimer, or higher noncovalent forms, spawning different models for its mechanism of action. We similarly show a large M(r)(app) by HPLC sieving chromatography, suggesting a tetrameric form. However, stoichiometry-ordered size (SOS) analysis of antibody-antigen complexes indicated only a single epitope per Tac molecule, compatible with a monomeric form. This larger M(r)(app) also conflicted with prior in vivo data showing rapid filtration of soluble Tac through the renal glomerulus that was not expected of a larger complex. Using different solvents, denaturants, and columns in the chromatography suggested that the elevated M(r)(app) values were an artifact of solute-column interactions, termed "ionic exclusion", rather than reflecting larger native structures. Analytical ultracentrifugation using a new type of analysis specific to glycoproteins demonstrated monomeric masses under all salt conditions with no tendency to form dimers or higher aggregates. Finally, circular dichroism spectroscopy showed no salt-dependent changes to suggest conformational alterations that might correlate with mobility changes on high pressure liquid chromatography. We conclude therefore that Tac is monomeric under physiologic conditions. Assessments of higher molecular weight for the purified soluble protein by other methods may be explained by the highly acidic nature of the molecule, which hampers matrix penetration with chromatographic media and by the high carbohydrate content and low partial specific volumes that accelerate the molecule in sedimentation media relative to pure protein standards.