Nuclear spectroscopic techniques for studying biological systems at ISOLDE

Abstract

Perturbed Angular Correlation of γ-rays technique (PAC) and β-decay NuclearMagnetic Resonance (β-NMR) are two very sensitive spectroscopic techniques,partly due to the use of radioactive isotopes. Nevertheless, they are not very frequently used among life-scientists due to both, the use of short-lived radioactiveisotopes requiring on-line production and the complexity of the experimental setups that are required. Conventional NMR on the other hand is one of the mostused and versatile techniques for studying the structure and dynamics of biochemical systems but lacks sensitivity. The three techniques belong to the family ofhyperfine interactions. They allow for measuring the interactions of the extranuclear electric and magnetic fields with the nuclear moments of the ensembleof nuclei under study. Thus, they can give insights to the local structure anddynamics for different time scales of the investigated systems.The first part of this work focuses on the PAC spectroscopy with heavy metalradioactive ions to explore the local electronic and molecular structure at thePAC probe site. Firstly, we focused on proving that a Cu isotope can be used tostudy the hyperfine interactions using the PAC technique. The 68mCu+ producedat ISOLDE-CERN was tested as the first Cu isotope and as a result its magneticdipole moment, µ, and the quadrupole moment, Q, was reported for the firsttime. This work was successful and clearly demonstrated that 68mCu/68Cu isa unique probe for hyperfine interactions in areas such as condensed and softmatter physics but it is unlikely that this isotope can be used for measurementsof NQIs of copper bound to biomolecules. Furthermore, 111mCd PAC experimentswere performed on a metalloregulatory protein found in two different bacteria, theCadC. We investigated the interactions between the 111mCd(II) heavy metal ionand the CadC protein. It is shown experimentally that two co-existing metal sitestructures in Listeria monocytogenes bacteria are in continuous rapid exchange fortemperatures higher than -196oC, the CdS4 and CdS3O. Furthermore, it is showniithat the binding site of CadC with Cd(II) and the dynamic exchange between thetwo sites is not affected by the binding of CadC with a DNA molecule. The second part of this thesis presents the numerous conventional 25Mg and 23NaNMR 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 25Mgand 23Na and the respective 1H NMR experimental spectra are presented. TheNMR measurements that constituted the reference for the β-detected NMR measurements were conducted later at ISOLDE-CERN. The third and most challenging part of this thesis was the design, building andput into operation of the experimental setup that could host liquid samples forapplying the β-detected NMR technique in biologically related liquids while usingthe radioactive beams from the ISOLDE targets. This work included the participation in building a new laser polarisation beamline at ISOLDE-CERN dedicatedto β-NMR experiments for studies in biochemistry and biophysics. The new vacuum compatible liquid handling system (LHS) tested off-line and on-line with ILsand radioactive beams was one of the main technical aspects if this work. Testsshowed 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 inchemistry and biochemistry. The results of the beamline commissioning and theNMR chemical shifts measured from different ILs are presented in this thesis.

Στην παρούσα διδακτορική διατριβή μελετήθηκαν τρεις διαφορετικές τεχνικές φασματοσκοπίας οι οποίες εφαρμόστηκαν σε υγρά, στερεά και βιολογικά δείγματα. Πιο συγκεκριμένα η διαταραγμένη γωνιακή συσχέτιση της τεχνικής γ-ακτίνων (PAC), ο πυρηνικός μαγνητικός συντονισμός και η τεχνική του β-NMR. Οι τεχνικές PAC και β-NMR είναι δύο πολύ ευαίσθητες φασματοσκοπικές τεχνικές, εν μέρει λόγω της χρήσης ραδιενεργών ισοτόπων. Ωστόσο, δεν χρησιμοποιούνται πολύ συχνά μεταξύ των βιοεπιστημόνων λόγω της χρήσης ραδιενεργών ισοτόπων βραχύβιας διάρκειαςπου απαιτούν on-line παραγωγή και την πολυπλοκότητα των πειραματικών ρυθμίσεων που απαιτούνται. Το συμβατικό NMR από την άλλη πλευρά είναι μια από τις πιο διαδεδομένες και ευέλικτες τεχνικές για τη μελέτη της δομής και της δυναμικής των βιοχημικών συστημάτων, αλλά στερείται ευαισθησίας. Οι τρεις τεχνικές ανήκουν στην οικογένεια των αλληλεπιδράσεων της υπέρλεπτης υφής . Επιτρέπουν τη μέτρηση των αλληλεπιδράσεων των εξωπυρηνικών ηλεκτρικών και μαγνητικών πεδίων με τις πυρηνικές ροπές του συνόλου υπό μελέτη πυρήνων. Έτσι, μπορούν να δώσουν πληροφορίες για την τοπική δομή καιδυναμική για διαφορετικές χρονικές κλίμακες των διερευνηθέντων συστημάτων.