An Iminodiacetic Acid Based Lanthanide Binding Tag for Paramagnetic Exchange NMR Spectroscopy

Abstract

When bound to proteins, paramagnetic lanthanide ions induce a range of effects that are observable by NMR spectroscopy, including pseudo-contact shifts (PCSs), paramagnetic relaxation enhancements (PREs), and residual dipolar couplings (RDCs).[1] These effects provide valuable constraints that can expedite protein structure refinement, [2]

Pseudocontact shifts in biomolecular NMR using paramagnetic metal tags

“…To reduce the size of the lanthanide tag even further, the iminodiacetic acid (IDA)- was found to position the lanthanide ion close to the carboxyl group of Asp32 located in the same a-helix [73]. The magnitudes of the Dc tensors observed for six different paramagnetic lanthanide ions followed the sequence observed for calbindin D9k (Fig.…”

“…Dc tensor for Dy 3+ , excellent correlations between experimental and back-calculated PCSs, and very good agreement between the Dc tensor and alignment tensor determined with Tb 3+ , indicating outstanding rigidity for such a small probe. Most interestingly, addition of an excess of a 1:1 mixture of diamagnetic La 3+ and paramagnetic Dy 3+ allowed the observation of exchange cross-peaks between the auto-peaks of the diamagnetic and paramagnetic protein, facilitating the assignment of the paramagnetic NMR spectrum [73], using an NMR experiment applied previously to measure PCSs in e186 [70]. Unfortunately, exchange crosspeaks have proven difficult to observe in other proteins (see Section 4.4.2).…”

FANTEN: a new web-based interface for the analysis of magnetic anisotropy-induced NMR data

“…The advantages offered by paramagnetic restraints are more and more exploited thanks to the development of paramagnetic tags that can be attached to diamagnetic proteins, so that PCSs and RDCs can be easily measured also for molecules without a natural metal binding site (Wöhnert et al 2003;Rodriguez-Castañeda et al 2006;Su et al 2006;Su et al 2008a;Keizers et al 2008;Su et al 2008b;Zhuang et al 2008;Häussinger et al 2009;Su and Otting 2010;Hass et al 2010;Man et al 2010;Das Gupta et al 2011;Liu et al 2012;Cerofolini et al 2013;Yagi et al 2013a;Gempf et al 2013;Kobashigawa et al 2012;Saio et al 2011;Watanabe et al 2010;Loh et al 2013;Swarbrick et al 2011aSwarbrick et al , 2011b.…”

New Lanthanide Tag for the Generation of Pseudocontact Shifts in DNA by Site-Specific Ligation to a Phosphorothioate Group

“…Uniformly 15 N-labeled samples of the ubiquitin mutant A28C were prepared and purified as described previously. 34 Samples were ligated with the C10 tag loaded with either Tb 3+ , Tm 3+ , Yb 3+ , or Y 3+ , and Dc tensors were determined from 15 N-HSQC spectra as described in the Supporting Information.…”