Classical molecular dynamics (MD) simulations, within the AMBER program package that runs entirely on a CUDA-enabled NVIDIA graphic processing unit (GPU), were employed to study with low computational cost and good quality of the sampling the dynamics of the methane-thiosulfonate spin label (MTSL) attached to the activation loop of the Aurora-A kinase. MD provided a wealth of information about the timescale of the different motional contributions to the overall dynamics of the spin label . These data were validated by multi-frequency continuous-wave electron paramagnetic resonance (EPR) measurements, that relying on the frequency dependence of the fast and slow motions of the spin probe were used to distinguish the fast internal motion of the spin label from slow protein tumbling. It was found that the activation loop oscillated between two conformational states separated by 7 Angstrom and the average structures obtained from the MD trajectories showed the MTSL exposed to the solvent and probing the C-lobe of the protein . The theoretical 9 and 94 GHz EPR spectra were calculated using configurations representing the interactions between MTSL and water and the tyrosine residue 208 in the C-lobe ; and the comparison with experimental EPR spectra revealed that fits successfully reproduced the experimental spectra in agreement with the MD results .
Classical molecular dynamics (MD) simulations, within the AMBER program package that runs entirely on a CUDA-enabled NVIDIA graphic processing unit (GPU), were employed to study with low computational cost the dynamics of the methane-thiosulfonate spin label (MTSL) attached to the activation loop of Aurora-A kinase. MD provided information about the conformational space of MTSL in the protein environment; an isotropic and uniform distribution of orientations of the spin label in space was found due to the large exposure of the activation loop to the solvent water. A hydrodynamic approach was employed to determine the rotational protein tumbling, while the internal motion of the spin probe was determined from 94 GHz measurements that reflect the fast motions . The theoretical 9 GHz EPR spectra were calculated using configurations representing interactions between MTSL and water and the tyrosine residue 208 in the C-lobe and the comparison with experimental EPR spectra produced fits that successfully reproduced the experimental spectra in agreement with the average structures obtained from the MD trajectories showing the MTSL exposed to the solvent and probing the C-lobe of the protein .
R
and good quality of the sampling the
the
D
the
D
a wealth of
R
timescale of
conformational space of MTSL in the protein environment; an isotropic and uniform distribution of orientations of
D
different motional contributions to the overall dynamics of the
R
. These data were validated by multi-frequency continuous-wave electron paramagnetic resonance (EPR) measurements, that relying on the frequency dependence of the fast and slow motions of the spin probe were used to distinguish the fast
in space was found due to the large exposure of the activation loop to the solvent water. A hydrodynamic approach was employed to determine the rotational protein tumbling, while the
R
label from slow protein tumbling. It was found that the activation loop oscillated between two conformational states separated by 7 Angstrom and the average structures obtained from the MD trajectories showed the MTSL exposed to the solvent and probing the C-lobe of the protein
probe was determined from 94 GHz measurements that reflect the fast motions
D
and 94
D
the
D
;
R
revealed that fits
produced fits that
R
MD results
average structures obtained from the MD trajectories showing the MTSL exposed to the solvent and probing the C-lobe of the protein