The optimization of the structural and magnetic properties of the polarizing agents has significantly contributed to the increase of the signal enhancement at 9 T and to the success of the technique. However, ssNMR/DNP still faces important disadvantages, such as non-optimal signal enhancement and a poor spectral resolution due to the low temperature of the experiments. Moreover, at high field (18 T), the gain in sensitivity is dramatically reduced in regards with experiments performed at 9T.
The proposed project will lead to the development of new polarizing agents and new methodologies that will make clearly beneficial the use of DNP at high field (18 T) and/or at higher temperature (above 200 K). Our initiatives will bring advances with efficient polarizing agents, a better understanding of the polarizing processes, and in the developments of ssNMR for the structural studies on challenging samples. The work will cover from the design and synthesis of new radicals, to EPR investigations at high field , DFT calculations and ssNMR DNP experiments at 9, 14 and 18 T and at various temperatures .
To meet these challenges, new families of polarizing agents will be introduced, such as strongly dipolar coupled dinitroxides, heterobiradicals, new narrow line radicals with pseudo-isotropic g values, new radicals for Overhauser Effect. Finally, these polarizing agents (PAs) will be exploited to record high-resolution high-temperature DNP enhanced NMR spectra on biological samples.
In a landmark paper, Griffin and co-workers have observed significant OE DNP in insulating solids . Capitalizing on this observation, we have investigated the MAS dependence of OE DNP using BDPA in dissolved in ortho-terphenyl (OTP. At 40 kHz MAS, an enhancement 105 was measured at 18.8 T. These results are not only the first data for OE DNP at very fast MAS, they also provide the highest enhancements obtained so far at a magnetic field strength of 18.8 T. Interestingly, we showed that the proton OE enhancement factors significantly increases with the MAS frequency. The contribution factors are close to one over the whole range of spinning speeds, indicating that, unlike CE nitroxide biradicals, BDPA-based formulation does not induce depolarization effects (i.e. signal reduction) upon fast sample spinning.
DNP represents a powerful way to overcome the sensitivity limitation of magic-angle spinning (MAS) NMR experiments. However, the resolution of the DNP NMR spectra of proteins is compromised bysevere line broadening associated with the necessity to perform experiments at cryogenic temperatures. We have shows that high quality DNP- enhanced NMR spectra of the Acinetobacterphage 205 (AP205) nucleocapsid can be obtained by combining high magnetic field (800 MHz) and fast MAS (40 kHz). This enables the assignment of aromatic resonances of the AP205 coat protein and its packaged RNA, which are not observed at room temperature, opening new possibilities for intermolecular interaction studies.
We recently focused our attention to the study of new families of polarizing agents suitable for DNP at very high field (18.8 T). In particular, we are currently investigating a series of new mixed radicals composed by a BDPA and a nitroxide moiety. The radicals show proton DNP enhancements of up to 100 at 9.4 T, and of up to 45 at 18.8 T, which surpasses the enhancements obtained by the most efficient binitroxides at this magnetic field.
New polarizing agents and methodologies enabling higher resolution DNP/ssNMR. These results open up new perspectives in the understanding of polarization process and transfer, and in the development of the technique, notably for structural studies of biomolecules.
3 présentations orales
Monsieur Olivier OUARI (Aix Marseille Université Institut de Chimie Radicalaire)
L'auteur de ce résumé est le coordinateur du projet, qui est responsable du contenu de ce résumé. L'ANR décline par conséquent toute responsabilité quant à son contenu.
ISA Institut des Sciences Analytiques
AMU Aix Marseille Université Institut de Chimie Radicalaire
Aide de l'ANR 335 180 euros
Début et durée du projet scientifique : septembre 2015 - 48 Mois