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The complexation of MgII with adenosine 5′‐triphosphate (ATP) is omnipresent in biochemical energy conversion, but is difficult to interrogate directly. Here we use the spin‐1/2 β‐emitter 31Mg to study MgII‐ATP complexation in 1‐ethyl‐3‐methylimidazolium acetate (EMIM‐Ac) solutions using β‐radiation‐detected nuclear magnetic resonance (β‐NMR). We demonstrate that (nuclear) spin‐polarized 31Mg, following ion‐implantation from an accelerator beamline into EMIM‐Ac, binds to ATP within its radioactive lifetime before depolarizing. The evolution of the spectra with solute concentration indicates that the implanted 31Mg initially bind to the solvent acetate anions, whereafter they undergo dynamic exchange and form either a mono‐ (31Mg‐ATP) or di‐nuclear (31MgMg‐ATP) complex. The chemical shift of 31Mg‐ATP is observed up‐field of 31MgMg‐ATP, in accord with quantum chemical calculations. These observations constitute a crucial advance towards using β‐NMR to probe chemistry and biochemistry in solution. [ABSTRACT FROM AUTHOR]

Since its development, the ionophore BAPTA (1,2‐bis(2‐aminophenoxy)‐ethane‐N,N,N',N'‐tetraacetic acid) has been used unchanged in calcium sensing applications. In this work we present a comprehensive experimental and theoretical study of novel alterations in the structure of BAPTA, with a focus on the systematic modification of the chain connecting the two aromatic rings of the molecule (denoted as "linker"). A bis‐(diethylamino)xantene fluorophore was also attached to the structures in a fixed position and the structure‐fluorescence response relationship of these molecules was investigated in addition. The effect of the linker's length, the number of oxygen atoms in this chain and even the removal of one of the rings was tested; these all proved to significantly alter the characteristics of the compounds. For example, it was found that the second aromatic ring of BAPTA is not essential for the turn‐on of the fluorescence. We also demonstrated that successful sensing can be realized even by replacing the chain with a single oxygen atom, which suggests the availability of a new calcium binding mode of the chelator. The reliable turn‐on characteristic, the steep Ca2+ fluorescence titration curve and the intense fluorescence emission combine to make this compound a prospective candidate as a calcium sensing molecular probe in diagnostic neurobiological applications. [ABSTRACT FROM AUTHOR]

Selectivity for monovalent metal ions is an important facet of the function of the metalloregulatory protein CueR. 111Ag perturbed angular correlation of γ‐rays (PAC) spectroscopy probes the metal site structure and the relaxation accompanying the instantaneous change from AgI to CdII upon 111Ag radioactive decay. That is, a change from AgI, which activates transcription, to CdII, which does not. In the frozen state (−196 °C) two nuclear quadrupole interactions (NQIs) are observed; one (NQI1) agrees well with two coordinating thiolates and an additional longer contact to the S77 backbone carbonyl, and the other (NQI2) reflects that CdII has attracted additional ligand(s). At 1 °C only NQI2 is observed, demonstrating that relaxation to this structure occurs within ≈10 ns of the decay of 111Ag. Thus, transformation from AgI to CdII rapidly disrupts the functional linear bis(thiolato)AgI metal site structure. This inherent metal site flexibility may be central to CueR function, leading to remodelling into a non‐functional structure upon binding of non‐cognate metal ions. In a broader perspective, 111Ag PAC spectroscopy may be applied to probe the flexibility of protein metal sites. [ABSTRACT FROM AUTHOR]

Intracellular CuI is controlled by the transcriptional regulator CueR, which effectively discriminates between monovalent and divalent metal ions. It is intriguing that HgII does not activate transcription, as bis‐thiolate metal sites exhibit high affinity for HgII. Here the binding of HgII to CueR and a truncated variant, ΔC7‐CueR, without the last 7 amino acids at the C‐terminus including a conserved CCHH motif is explored. ESI‐MS demonstrates that up to two HgII bind to CueR, while ΔC7‐CueR accommodates only one HgII. 199mHg PAC and UV absorption spectroscopy indicate HgS2 structure at both the functional and the CCHH metal site. However, at sub‐equimolar concentrations of HgII at pH 8.0, the metal binding site displays an equilibrium between HgS2 and HgS3, involving cysteines from both sites. We hypothesize that the C‐terminal CCHH motif provides auxiliary ligands that coordinate to HgII and thereby prevents activation of transcription. [ABSTRACT FROM AUTHOR]

Hg2+ and Cd2+ complexation of a short hexapeptide, Ac-DCSSCY-NH2 (DY), was studied by pH-potentiometry, UV and NMR spectroscopy and fluorimetry in aqueous solutions and the Hg2+-binding ability of the ligand was also described in an immobilized form, where the peptides were anchored to a hydrophilic resin. Hg2+ was demonstrated to form a 1 : 1 complex with the ligand even at pH = 2.0 while Cd2+ coordination by the peptide takes place only above pH ∼ 3.5. Both metal ions form bis-ligand complexes by the coordination of four Cys-thiolates at ligand excess above pH ∼ 5.5 (Cd2+) and 7.0 (Hg2+). Fluorescence studies demonstrated a Hg2+ induced concentration-dependent quenching of the Tyr fluorescence until a 1 : 1 Hg2+ : DY ratio. The fluorescence emission intensity decreases linearly with the increasing Hg2+ concentration in a range of over two orders of magnitude. The fact that this occurs even in the presence of 1.0 eq. of Cd2+ per ligand reflects a complete displacement of the latter metal ion by Hg2+ from its peptide-bound form. The immobilized peptide was also shown to bind Hg2+ very efficiently even from samples at pH = 2.0. However, the existence of lower affinity binding sites was also demonstrated by binding of more than 1.0 eq. of Hg2+ per immobilized DY molecule under Hg2+-excess conditions. Experiments performed with a mixture of four metal ions, Hg2+, Cd2+, Zn2+ and Ni2+, indicate that this molecular probe may potentially be used in Hg2+-sensing systems under acidic conditions for the measurement of μM range concentrations. [ABSTRACT FROM AUTHOR]