Conformational dynamics and distribution of nitroxide spin labels

Abstract

Long-range distance measurements based on paramagnetic relaxation enhancement (PRE) in NMR, quantification of surface water dynamics near biomacromolecules by Overhauser dynamic nuclear polarization (DNP) and sensitivity enhancement by solid-state DNP all depend on introducing paramagnetic species into an otherwise diamagnetic NMR sample. The species can be introduced by site-directed spin labeling, which offers precise control for positioning the label in the sequence of a biopolymer. However, internal flexibility of the spin label gives rise to dynamic processes that potentially influence PRE and DNP behavior and leads to a spatial distribution of the electron spin even in solid samples. Internal dynamics of spin labels and their static conformational distributions have been studied mainly by electron paramagnetic resonance spectroscopy and molecular dynamics simulations, with a large body of results for the most widely applied methanethiosulfonate spin label MTSL. These results are critically discussed in a unifying picture based on rotameric states of the group that carries the spin label. Deficiencies in our current understanding of dynamics and conformations of spin labeled groups and of their influence on NMR observables are highlighted and directions for further research suggested.

Structural basis for allosteric cross-talk between the asymmetric nucleotide binding sites of a heterodimeric ABC exporter

“…The experimental DEER distances agree with those simulated with a rotamer library approach (19) on the two available crystal structures, considering the 3 to 3.5Å accuracy achievable with this method (20) (Fig. 1 C-F Table S2), indicative of AMP-PNP binding and consequent allosteric effects in the intracellular region of the exporter.…”

Toward increased concentration sensitivity for continuous wave EPR investigations of spin-labeled biological macromolecules at high fields

“…Structure and structure-function relationships of macromolecules are areas of intense EPR effort [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16]. Coupled with site-directed spin-labeling (SDSL), EPR is oftentimes used to characterize protein and nucleic acid structures and dynamics, conformational changes, molecule folding, macromolecule complexes, and oligomeric structures [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15]17].…”

A combined EPR and MD simulation study of a nitroxyl spin label with restricted internal mobility sensitive to protein dynamics

“…A time scaling factor of ~1.5 applied to the MD trajectories was required to match the curves. It is common practice to remove global tumbling of the protein from the MD trajectories and re-introduce it by a BD approach (11,27,28). In order to achieve an accurate representation of the global tumbling of the protein molecule the BD trajectories for Mb were obtained by direct numerical solution of the Langevin equation (29,30) corresponding to the rotational diffusion described by three rotational diffusion coefficients Building up long total dynamical trajectories for EPR spectral simulations.…”

“…As has been highlighted and discussed in several recent reports (11,27,39,44) some rotameric states can be under represented in conventional MD trajectories and more advanced MD simulations, e.g. those employing multiple trajectories with different starting conformations (27) or parallel tempering simulations such as Replica Exchange MD (REMD) (34), would be required for adequate sampling to building up dynamical trajectories followed by the application of the kinetic modelling with either the Markov jump model (27) Figure 4 as bottom lines.…”

“…Although the inherent flexibility of R1, arising from five dihedral angles (Figure 1), allows the introduction of this spin label (SL) to many accessible sites of a protein it does not allow the unambiguous separation of side-chain motional dynamics from that of the protein. (9)(10)(11)(12)(13)(14) In fact, the rich conformational dynamics of the MTSL label and its distribution can significantly complicate both the analysis and interpretation of CW EPR spectra and long-range distance measurements in proteins using pulsed EPR techniques (11).…”