Binding: Metal ions

In addition to monitoring small molecule binding and protein-protein interactions, NMR has a long history in being used to study the binding of biologically important metal ions. Nature uses metals structurally and to provide chemical functionality and NMR can be used not only to demonstrate that metals are bound, but also to identify the binding site and define its geometry.

The most obvious NMR experiment to perform on a metal complex is to record a spectrum of the metal involved. Unfortunately, most biologically relevant metals do not produce good NMR spectra. The table below lists the NMR properties of some metals. Those with nuclear spin greater than ½ are highlighted in gray to indicate that they produce poor spectra with broad peaks due to their non zero quadropole moment that induces rapid T₂ relaxation.

Isotope	Nuclear Spin	γ (107 rad/T.s)	Natural Abundance (%)	Relative Sensitivity
1H	½	26.7519	99.98	100.00
13C	½	6.7283	1.108	1.59
15N	½	-2.712	0.365	0.104
23Na	3/2	7.0801	100.00	9.25
25Mg	5/2	-1.639	10.13	0.0267
31P	½	10.841	100.00	6.63
39K	3/2	1.2498	93.1	0.0508
43Ca	7/2	-1.8025	0.145	0.64
51V	7/2	7.0453	99.76	38
53Cr	3/2	-1.512	9.55	0.0903
55Mn	5/2	6.608	100.00	18
57Fe	½	0.8661	2.19	0.00337
59Co	7/2	6.317	100.00	28
63Cu	3/2	69.09	7.0974	0.0931
67Zn	5/2	1.6768	4.11	0.0285
95Mo	5/2	1.75	15.72	0.323
109Ag	½	-1.25	48.18	0.0101
113Cd	½	-5.995	12.26	1.09
119Sn	½	-10.0138	8.58	5.18
195Pt	½	5.768	33.7	0.994
199Hg	½	4.8154	16.84	0.567

To circumvent the poor spectra produced by most biologically relevant metals many researchers substitute the native metal ion with another, such as Cd²⁺, Hg⁺ or, less frequently, Ag⁺. The chemical shifts of bound ¹¹³Cd are characteristic of the metal's ligands and the geometry of the binding site. It is also possible to detect through bond interactions from the metal to the binding molecule and to measure three bond coupling constants and obtain torsion angles via the Karplus relationship. For tightly bound ions the native metal is removed by dialysis in the presence of a chelating agent, followed by reconstitution with the desired ion. If the metal ion is in exchange on the minute or second timescale then it can be replaced by equilibrium dialysis with a salt of the replacing ion.

Observation of metal ions requires the use of a broad band probe that can be tuned to the resonance frequency of the desired isotope. The Skaggs NMR Facility has a room temperature broad band probe, but it is not usually installed. The cryoprobes we normally use are optimised for detection of ¹H, ¹³C and ¹⁵N only.

Despite not being able to monitor most metal ions directly, it is possible to use NMR to follow changes to the neighbouring ¹H, ¹³C or ¹⁵N nuclei in the binding molecule. Metal ion titrations have the advantage that the titrant, in most cases, will not add extra, obscuring signals. Titrations with the metal can identify the binding site, the interacting atoms, the stoichiometry, and the dissociation constant.