NMR pseudocontact shifts in a symmetric protein homotrimer.

NMR pseudocontact shifts are a valuable tool for structural and functional studies of proteins. Protein multimers mediate key functional roles in biology, but methods for their study by pseudocontact shifts are so far not available. Paramagnetic tags attached to identical subunits in multimeric proteins cause a combined pseudocontact shift that cannot be described by the standard single-point model. Here, we report pseudocontact shifts generated simultaneously by three paramagnetic Tm-M7PyThiazole-DOTA tags to the trimeric molecular chaperone Skp and provide an approach for the analysis of this and related symmetric systems. The pseudocontact shifts were described by a "three-point" model, in which positions and parameters of the three paramagnetic tags were fitted. A good correlation between experimental data and predicted values was found, validating the approach. The study establishes that pseudocontact shifts can readily be applied to multimeric proteins, offering new perspectives for studies of large protein complexes by paramagnetic NMR spectroscopy.