Convenient Synthesis of Multifunctional EDTA‐Based Chiral Metal Chelates Substituted with an S‐Mesylcysteine

Abstract

We describe the synthetic route to ethylenediaminetetraacetic acid (EDTA) derivatives that can be attached to surface-exposed thiol functional groups of cysteine residues in proteins, via a methylthiosulfonate moiety that is connected in a stereochemically unique way to the C-1 carbon atom of EDTA. Such compounds can be used to align proteins in solution without the need to add liquid crystalline media, and are, therefore, of great interest for the NMR spectroscopic analysis of biomolecules. The binding constant for the paramagnetic tag to lanthanide ions was determined by measuring luminescence. For the Tb(+3)-ligand complex, a K(b) value of 6.5 x 10(17) M(-1) was obtained. This value is in excellent agreement with literature values for the related EDTA compound. In addition, it could be shown that there is no significant reduction in the luminescence intensity upon addition of a 10(4) excess of Ca2+ ions, indicating that this paramagnetic tag is compatible with buffers containing high concentrations of divalent alkaline earth ions.

Paramagnetic tagging of diamagnetic proteins for solution NMR

“…18 M range 18 and can therefore be used for the investigation of metal binding proteins. 19 They strongly align proteins at 800 MHz and lead to NH dipolar couplings exceeding 8 Hz.…”

Cys-Ph-TAHA: a lanthanide binding tag for RDC and PCS enhanced protein NMR

“…However, even less invasive is the attachment of a small chemically synthesized tag via a disulfide bridge to a single solvent exposed cysteine residue. Many tags based on organic ligands like EDTA (Gaponenko et al 2002; Ikegami et al 2004; Leonov et al 2005; Haberz et al 2006), DPA (Su et al 2008b; Man et al 2010), DOTA (Keizers et al 2007; Keizers et al 2008; Häussinger et al 2009), IDA (Swarbrick et al 2011a) or NTA (Swarbrick et al 2011b) have been developed to observe paramagnetic effects. So far, the largest RDCs obtained with paramagnetic tagging using a single cysteine residue have been reported with the DOTA-M8 and the IDA-based tag (Häussinger et al 2009; Swarbrick et al 2011a), while for two-point attachment even larger values have been measured by either using two NTA ligands (Swarbrick et al 2011b) or by connecting the tag via two disulfide bridges (CLaNP-5, Keizers et al 2008).…”

“…1) the following requirements were taken into account: complex stability for different temperatures and pH values, completeness of the tagging reaction and avoidance of the formation of diastereomeric complexes which would give rise to more than one set of NMR signals due to different tensors induced by different diastereomers of the tag (Ikegami et al 2004; Prudêncio et al 2004; Pintacuda et al 2004; Vlasie et al 2007). The latter was a severe problem with some of the first generation tags (Leonov et al 2005). In order to prevent these stereochemical problems we decided to synthesize a highly symmetric paramagnetic tag based on the TAHA chelator (Viguier et al 2001; Arslantas et al 2004).…”

Advanced NMR Techniques: Defining Carbohydrate Structures and Ligand–Receptor Interactions

“…Conformational studies of diamagnetic carbohydrates have been carried out by converting them to paramagnetic compounds through the attachment of an ethylenediaminetetraacetic acid (EDTA)-based paramagnetic tag, following the methodology already applied to proteins [40][41]. The use of this methodology has allowed for the characterization of disaccharides [34,41], trisaccharides [42], tetrasacharides [28], and very recently, a nonasaccharide [27].…”