Sub-THz specific relaxation times of hydrogen bond oscillations in E.coli thioredoxin. Molecular dynamics and statistical analysis.

Hydrogen bonds (H-bonds) in biological macromolecules are important for the molecular structure and functions. Since interactions via hydrogen bonds are weaker than covalent bonds, it can be expected that atomic movements involving H-bonds have low frequency vibrational modes. Sub-Terahertz (sub-THz) vibrational spectroscopy that combines measurements with molecular dynamics (MD) computational prediction has been demonstrated as a promising approach for biological molecule characterization. Multiple resonance absorption lines have been reported. The knowledge of relaxation times of atomic oscillations is critical for the successful application of THz spectroscopy for hydrogen bond characterization. The purpose of this work is to use atomic oscillations in the 0.35-0.7 THz range, found from molecular dynamic (MD) simulations of E.coli thioredoxin (2TRX), to study relaxation dynamics of two intra-molecular H-bonds, OH-N and OH-C. Two different complimentary techniques are used in this study, one is the analysis of the statistical distribution of relaxation time and dissipation factor values relevant to low frequency oscillations, and the second is the analysis of the autocorrelation function of low frequency quasi-periodic movements. By studying hydrogen bond atomic displacements, it was found that the atoms are involved in a number of collective oscillations, which are characterized by different relaxation time scales ranging from 2-3 ps to more than 150 ps. The existence of long lasting relaxation processes opens the possibility to directly observe and study H-bond vibrational modes in sub-THz absorption spectra of bio-molecules if measured with an appropriate spectral resolution. The results of measurements using a recently developed frequency domain spectroscopic sensor with a spectral resolution of 1 GHz confirm the MD analysis.

Molecular dynamics modeling of the sub-THz vibrational absorption of thioredoxin from E. coli.

Sub-terahertz (THz) vibrational modes of the protein thioredoxin in a water environment were simulated using molecular dynamics (MD) in order to find the conditions needed for simulation convergence, improve the correlation between experimental and simulated absorption frequencies, and ultimately enhance the predictive capabilities of computational modeling. Thioredoxin from E. coli was used as a model molecule for protocol development and to optimize the simulation parameters. The empirically parameterized software packages Amber 8 and 10 were used in this work. Using atomic trajectories from the constant energy and volume MD simulations, thioredoxin's sub-THz vibrational spectra and absorption coefficients were calculated in a quasi-harmonic approximation. An optimal production run length ~100 ps was found, in agreement with experimental data on thioredoxin relaxation dynamics. At the same time, a new procedure was developed for averaging correlation matrices of atomic coordinates in MD simulations. In particular, the open source package ptraj was edited to improve a matrix-analyzing function. Averaging only six matrices gave much more consistent results, with absorption peak intensities exceeding those from the individual spectra and a rather good correlation between simulated vibrational frequencies and experimental data.

Presynaptic Ca2+ buffers control the strength of a fast post-tetanic hyperpolarization mediated by the alpha3 Na(+)/K(+)-ATPase.

The excitability of CNS presynaptic terminals after a tetanic burst of action potentials is important for synaptic plasticity. The mechanisms that regulate excitability, however, are not well understood. Using direct recordings from the rat calyx of Held terminal, we found that a fast Na(+)/K(+)-ATPase (NKA)-mediated post-tetanic hyperpolarization (PTH) controls the probability and precision of subsequent firing. Notably, increasing the concentration of internal Ca(2+) buffers or decreasing Ca(2+) influx led to larger PTH amplitudes, indicating that an increase in [Ca(2+)](i) regulates PTH via inhibition of NKAs. The characterization for the first time of a presynaptic NKA pump current, combined with immunofluorescence staining, identified the alpha3-NKA isoform on calyx terminals. Accordingly, the increased ability of the calyx to faithfully fire during a high-frequency train as it matures is paralleled by a larger expression of alpha3-NKA during development. We propose that this newly discovered Ca(2+) dependence of PTH is important in the post-burst excitability of nerve terminals.