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Long-Lived States in Hyperpolarized Deuterated Methyl Groups Reveal Weak Binding of Small Molecules to Proteins

J. Phys. Chem. Lett. 2019, 10, 1523−1529 


We introduce a method for the detection of weak interactions of small molecules such as metabolites or medicaments that contain deuterated methyl groups with proteins in solution. The technique relies on long-lived imbalances of spin state populations, which are generated by dissolution dynamic nuclear polarization (D-DNP) and feature lifetimes that depend on the frequency of internal rotation of deuterated methyl groups. We demonstrate the technique for interactions between deuterated dimethyl sulfoxide (DMSO-d6) and bovine serum albumin (BSA) or trypsin, where the methyl group rotation is slowed down upon protein binding, which causes a marked reduction in the lifetime of the population imbalances. 

Time-Resolved Protein Side-Chain Motions Unraveled by High- Resolution Relaxometry and Molecular Dynamics Simulations

Journal of the American Chemical SocietyVolume 140, p. 13456 - 13465, 2018


Motions of proteins are essential for the performance of their functions. Aliphatic protein side chains and their motions play critical roles in protein interactions: for recognition and binding of partner molecules at the surface or serving as an entropy reservoir within the hydrophobic core. Here, we present a new NMR method based on highresolution relaxometry and high-fi eld relaxation to determine quantitatively both motional amplitudes and time scales of methyl-bearing side chains in the picosecond-to-nanosecond range. We detect a wide variety of motions in isoleucine side chains in the protein ubiquitin. We unambiguously identify slow motions in the low nanosecond range, which, in conjunction with molecular dynamics computer simulations, could be assigned to transitions between rotamers. Our approach provides unmatched detailed insight into the motions of aliphatic side chains in proteins and provides a better understanding of the nature and functional role of protein side-chain motions.


Dramatic Decrease in CEST Measurement Times Using Multi-Site Excitation

ChemPhysChem 2018, 19, 1707-1710


Chemical exchange saturation transfer (CEST) has recently evolved into a powerful approach for studying sparsely populated, “invisible” protein states in slow exchange with a major, visible conformer. Central to the technique is the use of a weak, highly selective radio‐frequency field that is applied at different frequency offsets in successive experiments, “searching” for minor state resonances. The recording of CEST profiles with enough points to ensure coverage of the entire spectrum at sufficient resolution can be time‐consuming, especially for applications that require high static magnetic fields or when small chemical shift differences between exchanging states must be quantified. Here, we show – with applications involving 15N CEST – that the process can be significantly accelerated by using a multi‐frequency irradiation scheme, leading in some applications to an order of magnitude savings in measurement time.


Sample Ripening through Nanophase Separation Influences the Performance of Dynamic Nuclear Polarization

Angew. Chem. Int. Ed. 2018, 57, 5171 –5175


Mixtures of water and glycerol provide popular matrices for low-temperature spectroscopy of vitrified samples. However, they involve counterintuitive physicochemical properties, such as spontaneous nanoscopic phase separations (NPS) in solutions that appear macroscopically homogeneous. We demonstrate that such phenomena can substantially influence the efficiency of dynamic nuclear polarization (DNP) by factors up to 20% by causing fluctuations in local concentrations of polarization agents (radicals). Thus, a spontaneous NPS of water/glycerol mixtures that takes place on time scales on the order of 30–60 min results in a confinement of polarization agents in nanoscopic water-rich vesicles, which in return affects the DNP. Such effects were found for three common polarization agents, TEMPOL, AMUPol and Trityl.

Graftable SCoMPIs enable the labeling and X-ray fluorescence imaging of proteins

Chem. Sci.2018, 9, 4483-4487


Bio-imaging techniques alternative to fluorescence microscopy are gaining increasing interest as complementary tools to visualize and analyze biological systems. Among them, X-ray fluorescence microspectroscopy provides information on the local content and distribution of heavy elements (Z > 14) in cells or biological samples. In this context, similar tools to those developed for fluorescence microscopy are desired, including chemical probes or tags. In this work, we study rhenium complexes as a convenient and sensitive probe for X-ray fluorescence microspectroscopy. We demonstrate their ability to label and sense exogenously incubated or endogenous proteins inside cells.