Nuclear Spectroscopic Techniques for studying Biological Systems at ISOLDE

Perturbed Angular Correlation of γ-rays technique (PAC) and $\beta$-decay Nuclear Magnetic Resonance ($\beta$-NMR) are two very sensitive spectroscopic techniques, partly due to the use of radioactive isotopes. Nevertheless, they are not very fre- quently used among life-scientists due to both, the use of short-lived radioactive isotopes requiring on-line production and the complexity of the experimental se- tups that are required. Conventional NMR on the other hand is one of the most used and versatile techniques for studying the structure and dynamics of biochem- ical systems but lacks sensitivity. The three techniques belong to the family of hyperfine interactions. They allow for measuring the interactions of the extra- nuclear electric and magnetic fields with the nuclear moments of the ensemble of nuclei under study. Thus, they can give insights to the local structure and dynamics for different time scales of the investigated systems. The first part of this work focuses on the PAC spectroscopy with heavy metal radioactive ions to explore the local electronic and molecular structure at the PAC probe site. Firstly, we focused on proving that a Cu isotope can be used to study the hyperfine interactions using the PAC technique. The $^{68m}$Cu$^+$ produced at ISOLDE-CERN was tested as the first Cu isotope and as a result its magnetic dipole moment, $\mu$, and the quadrupole moment, $Q$, was reported for the first time. This work was successful and clearly demonstrated that $^{68m}$Cu/$^{68}$Cu is a unique probe for hyperfine interactions in areas such as condensed and soft matter physics but it is unlikely that this isotope can be used for measurements of NQIs of copper bound to biomolecules. Furthermore, $^{111m}$Cd PAC experiments were performed on a metalloregulatory protein found in two different bacteria, the CadC. We investigated the interactions between the $^{111m}$Cd(II) heavy metal ion and the CadC protein. It is shown experimentally that two co-existing metal site structures in Listeria monocytogenes bacteria are in continuous rapid exchange for temperatures higher than -196$^{\circ}$C, the CdS$_4$ and CdS$_3$O. Furthermore, it is shownthat the binding site of CadC with Cd(II) and the dynamic exchange between the two sites is not affected by the binding of CadC with a DNA molecule. The second part of this thesis presents the numerous conventional $^{25}$Mg and $^{23}$Na NMR measurements in ionic liquids, that took place at the University of Copenhagen with a 11.74 T magnet. A thorough analysis and comparison of the $^{25}$Mg and $^{23}$Na and the respective $^1$H NMR experimental spectra are presented. The NMR measurements that constituted the reference for the $\beta$-detected NMR measurements were conducted later at ISOLDE-CERN. The third and most challenging part of this thesis was the design, building and put into operation of the experimental setup that could host liquid samples for applying the $\beta$-detected NMR technique in biologically related liquids while using the radioactive beams from the ISOLDE targets. This work included the participation in building a new laser polarisation beamline at ISOLDE-CERN dedicated to $\beta$-NMR experiments for studies in biochemistry and biophysics. The new vacuum compatible liquid handling system (LHS) tested off-line and on-line with ILs and radioactive beams was one of the main technical aspects if this work. Tests showed that the system is reliable and allowed for measuring the first NMR chemical shifts from non-aqueous, ionic liquid hosts paving the way to applications in chemistry and biochemistry. The results of the beamline commissioning and the NMR chemical shifts measured from different ILs are presented in this thesis.