Solid-state nuclear magnetic resonance of rhodopsin and its photointermediates

Photoisomerization of the membrane-bound light receptor protein rhodopsin leadsto a highly energetic species called bathorhodopsin, which is stable at temperaturesbelow 125 K. Bathorhodopsin stores about 2/3 of the absorbed photon energy butthe mechanisms with which this energy is stored is not completely understood. Anew insight into these mechanisms by means of low-temperature solid-state NMRis both subject and aim of this Ph.D. thesis. The issue of the energy storage hasbeen investigated by a solid state magic angle spinning technique which combinesmodern symmetry-based recoupling techniques with in situ cooling of the sample.Production of bathorhodopsin is also done in situ in a customized NMR probe.Three kind of experiments are discussed: chemical shift, distance and torsionalangle measurements. The first kind of experiments led to carbon chemical shiftsvalues for almost all the carbons along the retinylidene chain of the retinal chromophoreof bathorhodopsin. Our measurements show a significant perturbations ofthe 13C chemical shifts in bathorhodopsin which is interpreted in terms of chargedelocalization along the chain and therefore indicates a participation of an electrostaticmechanism to the energy storage. This is at variance with an earlier solidstate NMR study where only minor perturbations of the electronic structure in theisomerized retinylidene chain were observed. We believe that these data incorrectlyrefer to bathorhodopsin because of the incorrect conditions of temperature and illuminationapplied. To sample for other local mechanisms that may contribute to theenergy storage, the C-C distance of the last two carbons of the retinylidene chain,at the link with the protein opsin, was also measured but no significant differenceswith rhodopsin have been found. Finally, the H-C=C-H torsional angle at the doublebound where the isomerization takes place was measured in a double-quantumheteronuclear local field spectroscopy (2Q-HLF) experiment. Results indicate adeviation from planarity of at least 40? about this double bond in bathorhodopsinsuggesting an unquantified amount of torsional strain acting as a further energystorage mechanism. In addition to these very interesting results, this thesis reportsmethods, equipment and procedures ready to be used for the study of other similarlight-triggered processes

Unique Group 1 cations stabilised by homoleptic neutral phosphine coordination

Homoleptic coordination of the neutral diphosphines Me2P(CH2)2PMe2 and o-C6H4(PMe2)2 to the hard Li+ and Na+ cations is achieved using Li[Al{OC(CF3)3}4] and Na[B{3,5-(CF3)2-C6H3}4] as ‘naked’ cation sources. Crystallographic, solid state and solution multinuclear NMR studies confirm distorted octahedral coordination solely via three chelating diphosphines in these unique species

Rare neutral diphosphine complexes of scandium(III) and yttrium(III) halides

Reaction of Me2PCH2CH2PMe2 or o-C6H4(PMe2)2(L?L) with a suspension of ScI3 or YI3 in MeCN solution under rigorously anhydrous and oxygen-free conditions produced the highly unusual complexes [ScI3(L?L)2], [YI3(Me2PCH2CH2PMe2)2], and [YI3{o-C6H4(PMe2)2}2MeCN]. X-ray crystal structures reveal that the scandium complexes adopt seven-coordinate, pentagonal-bipyramidal geometries with chelating diphosphines, while the eight-coordinate[YI3{o-C6H4(PMe2)2}2MeCN] is dodecahedral. The complexes were characterized by microanalysis and IR and multinuclear NMR spectroscopy. Solid-state NMR data (45Sc, 89Y, 31P) and variable-temperature solution NMR data (1H, 31P{1H}, 45Sc) are presented and compared, leading to the conclusion that the same species are present in both the solid state and CH2Cl2 solution. Attempts to prepare complexes with other scandium halides and with aryl diphosphines and o-C6H4(AsMe2)2 are briefly described

14N overtone NMR under MAS: signal enhancement using cross-polarization methods

Polarization transfer methods are widely adopted for the purpose of correlating different nuclear species as well as to achieve signal enhancement. Polarization transfer from 1H to the 14N overtone transition (Δm=2) can be achieved using cross polarization methods under magic-angle spinning conditions, where spin locks of the order of several milliseconds can be obtained on common bio-solids (α-glycine and N-acetylvaline). Signal enhancement factors up to 4.4 per scan, can be achieved under favorable conditions, despite MHz-sized quadrupolar interaction. Moreover, we present a detailed theoretical treatment and accurate numerical simulations which are in excellent agreement the unusual experimental matching conditions observed for cross-polarization to 14N overtone

Quantitative analysis of Q2 14N quadrupolar coupling using 1H detected 14N solid-state NMR†

Magic-angle spinning solid-state NMR is increasingly utilized to study the naturally abundant, spin-1 nucleus 14N, providing insights into the structure and dynamics of biological and organic molecules. In particular, the characterisation of 14N sites using indirect detection has proven useful for complex molecules, where the ‘spy’ nucleus provides enhanced sensitivity and resolution. Here we exploit the sensitivity of proton detection, to indirectly characterise 14N sites using a moderate rf field to generate coherence between the 1H and 14N at moderate and fast-magic-angle spinning frequencies. Efficient numerical simulations have been developed that have allowed us to quantitatively analyse the resulting 14N lineshapes to determine both the size and asymmetry of the quadrupolar interaction. Exploiting only naturally occurring abundant isotopes will aid the analysis of materials with the need to resort to isotope labelling, whilst providing additional insights into the structure and dynamics that the characterisation of the quadrupolar interaction affords

Rare Neutral Diphosphine Complexes of Scandium(III) and Yttrium(III) Halides

Reaction of Me₂PCH₂CH₂PMe₂ or <i>o</i>-C₆H₄(PMe₂)₂ (L–L) with a suspension of ScI₃ or YI₃ in MeCN solution under rigorously anhydrous and oxygen-free conditions produced the highly unusual complexes [ScI₃(L–L)₂], [YI₃(Me₂PCH₂CH₂PMe₂)₂], and [YI₃{<i>o</i>-C₆H₄(PMe₂)₂}₂MeCN]. X-ray crystal structures reveal that the scandium complexes adopt seven-coordinate, pentagonal-bipyramidal geometries with chelating diphosphines, while the eight-coordinate [YI₃{<i>o</i>-C₆H₄(PMe₂)₂}₂MeCN] is dodecahedral. The complexes were characterized by microanalysis and IR and multinuclear NMR spectroscopy. Solid-state NMR data (⁴⁵Sc, ⁸⁹Y, ³¹P) and variable-temperature solution NMR data (¹H, ³¹P­{¹H}, ⁴⁵Sc) are presented and compared, leading to the conclusion that the same species are present in both the solid state and CH₂Cl₂ solution. Attempts to prepare complexes with other scandium halides and with aryl diphosphines and <i>o</i>-C₆H₄(AsMe₂)₂ are briefly described

Syntheses of 13C2-labelled 11Z-retinals

To enable solid-state NMR investigations of the rhodopsin chromophore and its photointermediates, a series of 11Z-retinal isotopomers have been synthesised containing pairs of adjacent 13C labels at C9/C10, C10/C11 or C11/C12, respectively. The C9 labelled carbon atom was introduced through the Heck reaction of a 13C-labelled Weinreb acrylamide derivative, and the label at the C12 position derived from a 13C-containing ethoxy Bestmann–Ohira reagent. The 13C labels at C10 and C11 were introduced through the reaction of ?-ionone with labelled triethyl phosphonoacetat

Rare Neutral Diphosphine Complexes of Scandium(III) and Yttrium(III) Halides

Reaction of Me₂PCH₂CH₂PMe₂ or <i>o</i>-C₆H₄(PMe₂)₂ (L–L) with a suspension of ScI₃ or YI₃ in MeCN solution under rigorously anhydrous and oxygen-free conditions produced the highly unusual complexes [ScI₃(L–L)₂], [YI₃(Me₂PCH₂CH₂PMe₂)₂], and [YI₃{<i>o</i>-C₆H₄(PMe₂)₂}₂MeCN]. X-ray crystal structures reveal that the scandium complexes adopt seven-coordinate, pentagonal-bipyramidal geometries with chelating diphosphines, while the eight-coordinate [YI₃{<i>o</i>-C₆H₄(PMe₂)₂}₂MeCN] is dodecahedral. The complexes were characterized by microanalysis and IR and multinuclear NMR spectroscopy. Solid-state NMR data (⁴⁵Sc, ⁸⁹Y, ³¹P) and variable-temperature solution NMR data (¹H, ³¹P­{¹H}, ⁴⁵Sc) are presented and compared, leading to the conclusion that the same species are present in both the solid state and CH₂Cl₂ solution. Attempts to prepare complexes with other scandium halides and with aryl diphosphines and <i>o</i>-C₆H₄(AsMe₂)₂ are briefly described

Fine structure in the solution state 13C-NMR spectrum of C60 and its endofullerene derivatives

The 13C NMR spectrum of fullerene C60 in solution displays two small “side peaks" on the shielding side of the main 13C peak, with integrated intensities of 1.63% and 0.81% of the main peak. The two side peaks are shifted by -12.6 ppb and -20.0 ppb with respect to the main peak. The side peaks are also observed in the 13C NMR spectra of endofullerenes, but with slightly different shifts relative to the main peak. We ascribe the small additional peaks to minor isotopomers of C60 containing two adjacent 13C nuclei. The shifts of the additional peaks are due to a secondary isotope shift of the 13C resonance caused by the substitution of a 12C neighbour by 13C. Two peaks are observed since the C60 structure contains two different classes of carbon-carbon bonds with different vibrational characteristics. The 2:1 ratio of the side peak intensities is consistent with the known structure of C60. The origin and intensities of the 13C side peaks are discussed, together with an analysis of the 13C solution NMR spectrum of a 13C-enriched sample of C60, which displays a relatively broad 13C NMR peak due to a statistical distribution of 13C isotopes. The spectrum of 13C-enriched C60 is analyzed by a Monte Carlo simulation technique, using a theorem for the second moment of the NMR spectrum generated by J-coupled spin clusters.<br/

An internuclear J-coupling of ³He induced by molecular confinement

The solution-state 13C NMR spectrum of the endofullerene 3He@C60 displays a doublet structure due to a J-coupling of magnitude 77.5 ± 0.2 mHz at 340K between the 3He nucleus and a 13C nucleus of the enclosing carbon surface. The J-coupling increases in magnitude with increasing temperature. Quantum chemistry calculations successfully predict the approximate magnitude of the coupling. This observation shows that the mutual proximity of molecular or atomic species is sufficient to induce a finite scalar nuclear spin-spin coupling, providing that translational motion is restricted by confinement. The phenomenon may have applications to the study of surface interactions and to mechanically bound species