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In this paper, the rotational correlation time for Human Serum Albumin and b-Amyloid1-16 was tried to be determined through Fluorescence Anisotropy and Perturbed Angular Correlation. While other types of spectroscopies, such as Ultraviolet, Optical Rotation, Thermal Protein Unfolding, and Decay Time, were used for experimental specifications. Through fluorescence anisotropy, decay time measurements, and theoretically calculated rotational correlation times, it was determined that it was not possible to measure the rotational correlation time for human serum albumin using this method. However, the measured rotational correlation time through perturbed angular correlation in water was close to the theoretical rotational correlation time for human serum albumin in water. These measurements suggested that human serum albumin will bind with mercury in the solution in a distorted linear structure. It was found to be possible to measure the rotational correlation time for b-Amyloid1-16 and determine whether the measurements were made on a monomer, dimer, or trimer. However, it has not been able to reproduce the measurements made priorly by Mikhail Baibakov.

With this work we first test various DFT functionals against CCSD(T) for calculation of EFGs at the position of Cd(II) in a very small model system, Cd(SCH3)2. Moreover, the available basis sets in ADF are tested in terms of basis set convergence, and the effect of including relativistic effects using the scalar relativistic and spin orbit ZORA hamiltonians is explored. The results indicate that an error of up to around 10 % on the calculated EFG may be expected using spin-orbit ZORA and the BHandH functional with a locally dense basis set. Next, this method was applied to model systems of the CueR protein, aiming to interpret 111Ag-PAC spectroscopic data. Note that 111Ag decays to 111Cd on which the PAC data are recorded. Surprisingly, model systems truncated - as is often done - at the first C-C bond from the central Cd(II) are inadequate in size, and larger model systems must be employed to achieve reliable EFG calculations. The calculated EFGs agree well with experimental PAC data, and indicate that shortly after the nuclear decay the structure relaxes from linear two-coordinate AgS2 in the native protein, to a structure (or structures) where Cd(II) recruits additional ligands such as backbone carbonyl oxygens to achieve higher coordination number(s).

In nature, thiolate-based systems are the primary targets of divalent mercury (Hg-II) toxicity. The formation of Hg(Cys)(x) cores in catalytic and structural protein centers mediates mercury's toxic effects and ultimately leads to cellular damage. Multiple studies have revealed distinct Hg-II-thiolate coordination preferences, among which linear Hg-II complexes are the most commonly observed in solution at physiological pH. Trigonal or tetrahedral geometries are formed at basic pH or in tight intraprotein Cys-rich metal sites. So far, no interprotein tetrahedral Hg-II complex formed at neutral pH has been reported. Rad50 protein is a part of the multiprotein MRN complex, a major player in DNA damage-repair processes. Its central region consists of a conserved CXXC motif that enables dimerization of two Rad50 molecules by coordinating Zn-II. Dimerized motifs form a unique interprotein zinc hook domain (Hk) that is critical for the biological activity of the MRN. Using a series of length-differentiated peptide models of the Pyrococcus furiosus zinc hook domain, we investigated its interaction with Hg-II. Using UV-Vis, CD, PAC, and Hg-199 NMR spectroscopies as well as anisotropy decay, we discovered that all Rad50 fragments preferentially form homodimeric Hg(Hk)(2) species with a distorted tetrahedral HgS4 coordination environment at physiological pH; this is the first example of an interprotein mercury site displaying tetrahedral geometry in solution. At higher Hg-II content, monomeric HgHk complexes with linear geometry are formed. The Hg(Cys)(4) core of Rad50 is extremely stable and does not compete with cyanides, NAC, or DTT. Applying ITC, we found that the stability constant of the Rad50 Hg(Hk)(2) complex is approximately three orders of magnitude higher than those of the strongest Hg-II complexes known to date.

The transcriptional regulator CueR is activated by the binding of Cu-I, Ag-I, or Au-I to two cysteinates in a near-linear fashion. The C-terminal CCHHRAG sequence in Escherichia coli CueR present potential additional metal binding ligands and here we explore the effect of deleting this fragment on the binding of Ag-I to CueR. CD spectroscopic and ESI-MS data indicate that the high Ag-I-binding affinity of WT-CueR is significantly reduced in Delta 7C-CueR.([111) Ag PAC spectroscopy demonstrates that the WT-CueR metal site structure (AgS2) is conserved, but less populated in the truncated variant. Thus, the function of the C-terminal fragment may be to stabilize the two-coordinate metal site for cognate monovalent metal ions. In a broader perspective this is an example of residues beyond the second coordination sphere affecting metal site physicochemical properties while leaving the structure unperturbed.

In nature, thiolate-based systems are the primary targets of divalent mercury (HgII ) toxicity. The formation of Hg(Cys)x cores in catalytic and structural protein centers mediates mercury's toxic effects and ultimately leads to cellular damage. Multiple studies have revealed distinct HgII -thiolate coordination preferences, among which linear HgII complexes are the most commonly observed in solution at physiological pH. Trigonal or tetrahedral geometries are formed at basic pH or in tight intraprotein Cys-rich metal sites. So far, no interprotein tetrahedral HgII complex formed at neutral pH has been reported. Rad50 protein is a part of the multiprotein MRN complex, a major player in DNA damage-repair processes. Its central region consists of a conserved CXXC motif that enables dimerization of two Rad50 molecules by coordinating ZnII . Dimerized motifs form a unique interprotein zinc hook domain (Hk) that is critical for the biological activity of the MRN. Using a series of length-differentiated peptide models of the Pyrococcus furiosus zinc hook domain, we investigated its interaction with HgII . Using UV-Vis, CD, PAC, and 199 Hg NMR spectroscopies as well as anisotropy decay, we discovered that all Rad50 fragments preferentially form homodimeric Hg(Hk)2 species with a distorted tetrahedral HgS4 coordination environment at physiological pH; this is the first example of an interprotein mercury site displaying tetrahedral geometry in solution. At higher HgII content, monomeric HgHk complexes with linear geometry are formed. The Hg(Cys)4 core of Rad50 is extremely stable and does not compete with cyanides, NAC, or DTT. Applying ITC, we found that the stability constant of the Rad50 Hg(Hk)2 complex is approximately three orders of magnitude higher than those of the strongest HgII complexes known to date.

Accurate nuclear quadrupole moment values are essential as benchmarks for nuclear structure models and for the interpretation of experimentally determined nuclear quadrupole interactions in terms of electronic and molecular structure. Here we present a novel route to such data by combining perturbed γ-γ angular correlation measurements on free small linear molecules, realized for the first time within this work, with state-of-the-art ab initio electronic structure calculations of the electric field gradient at the probe site. This approach, also feasible for a series of other cases, is applied to Hg and Cd halides, resulting in Q(199Hg,5/2-) = +0.674(17) b and Q(111Cd,5/2+) = +0.664(7) b.

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.