Periodic Reporting for period 2 - BIOVIB (Electric Interactions and Structural Dynamics of Hydrated Biomolecules Mapped by Ultrafast Vibrational Probes)
Période du rapport: 2020-11-01 au 2022-04-30
Biomolecules exist in a liquid environment of water molecules and ions. Water molecules possess an electric dipole moment and ions are charged, leading to strong electric interactions within the liquid and with embedded biomolecules. Their structure and biological function are, thus, strongly influenced by the fluctuating water shell and the ion distribution. At the molecular level, such interactions are barely understood; the electric field strengths, spatial range and interplay with other interactions are mainly unknown. Insight in this fundamental problem is important for designing and synthesizing biomolecular systems with specific and tailored functional properties for applications in bioengineering, pharmacology, and other areas of societal relevance.
Going far beyond existing methods, this project introduces a novel scientific paradigm for a direct time-resolved mapping of molecular electric forces on sub-nanometer length scales and at the genuine terahertz (THz) fluctuation frequencies of biomolecules embedded in water. Femtosecond vibrational excitations of DNA and RNA at the interface to the water shell act as noninvasive probes of charge dynamics and local electric fields. Key objectives are quantitative measurements and simulations of dynamic electric fields acting on prototypical DNA, RNA, and membrane structures, another the analysis of ion geometries around charged biomolecules and their role in stabilizing structure. This includes a critical assessment of existing concepts for describing electrostatics at the molecular scale and a clarification of the particular role of the water environment.
Going far beyond existing methods, this project introduces a novel scientific paradigm for a direct time-resolved mapping of molecular electric forces on sub-nanometer length scales and at the genuine terahertz (THz) fluctuation frequencies of biomolecules embedded in water. Femtosecond vibrational excitations of DNA and RNA at the interface to the water shell act as noninvasive probes of charge dynamics and local electric fields. Key objectives are quantitative measurements and simulations of dynamic electric fields acting on prototypical DNA, RNA, and membrane structures, another the analysis of ion geometries around charged biomolecules and their role in stabilizing structure. This includes a critical assessment of existing concepts for describing electrostatics at the molecular scale and a clarification of the particular role of the water environment.
The work done in the reporting period followed three main directions, the implementation of new methods for THz spectroscopy of biomolecules in a water environment, the investigation of the response of water to high external THz fields, and in-depth studies of the interactions of RNA with water and ions by two-dimensional infrared (2D-IR) spectroscopy.
Laser-driven THz sources provide picosecond electric field transients with a peak amplitude of up to 2 MV/cm. Interaction of the THz pulses with metallic antenna structures allows for electric field enhancement by a factor of 3 to 5, as was demonstrated in measurements and simulations. For time-resolved experiments with THz pump and mid-infrared probe pulses, a sample geometry including the antennas and a thin liquid film was developed.
The nonlinear response of liquid water to strong THz fields was studied by 2D-THz spectroscopy in the nonperturbative regime of light-matter interaction. In presence of a THz electric field beyond 500 kV/cm, the generation of solvated electrons by ionization of water molecules was observed for the first time. Tunnel ionization of water molecules is induced by the strong fluctuation electric field from water dipoles reaching peak values on the order of 100 MV/cm, while the external THz field serves for a spatial transport of the released electrons away from the parent ion. The electron eventually localizes at a new site in the liquid, a process occurring on a time scale of a few picoseconds.
The solvated electron is electrically coupled to a large number of water molecules in its environment. This many-body interactions manifests in new types of collective excitations of the ensemble. Femtosecond optical pump - THz probe experiments provide the first evidence for polarons in water which are hybrid excitations of the electron and librational motions of water molecules. The polaron displays coherent oscillations in a picosecond time range and with a frequency determined by the electron concentration. The surprisingly weak damping of polaron oscillations originates from their longitudinal character and the weak coupling to other excitations of the liquid at such low frequency.
The water geometries at the surface of double-stranded helical RNA and transfer RNA (tRNA) were studied by 2D-IR spectroscopy of vibrations of the RNA backbone which serve as noninvasive probes of local interactions. The 2D-IR spectra of the asymmetric phosphate stretching vibration of tRNA reveal three prototypical hydration motifs (see Figure), a local hydration shell consisting of six water molecules forming hydrogen bonds with the free phosphate oxygens, an ordered water geometry in which individual water molecules bridge neighboring phosphate groups, and hydrated contact pairs of phosphate groups with counterions. The spectroscopic observations were analyzed by in-depth theoretical calculations and molecular dynamics simulations. RNA melting leads to a decrease of ordered structures in favor of local hydration shells. Librational water motions lead to structural fluctuations on a time scale of 300 fs while hydrogen bond lifetimes and reordering times of ions are much longer.
The work on RNA was complemented by studies of phosphate/magnesium interactions with the help of the model system dimethylphosphate. Linear and 2D-IR spectroscopy allowed for establishing clear spectroscopic signatures of contact ion pairs and underlined the particularly strong electrostatic interaction of the ions embedded in a comparably rigid water geometry. This behavior is in contrast to ion pairs with sodium and calcium ions. Currently, this work is being extended to transfer RNA structures embedded in an atmosphere of magnesium ions.
Laser-driven THz sources provide picosecond electric field transients with a peak amplitude of up to 2 MV/cm. Interaction of the THz pulses with metallic antenna structures allows for electric field enhancement by a factor of 3 to 5, as was demonstrated in measurements and simulations. For time-resolved experiments with THz pump and mid-infrared probe pulses, a sample geometry including the antennas and a thin liquid film was developed.
The nonlinear response of liquid water to strong THz fields was studied by 2D-THz spectroscopy in the nonperturbative regime of light-matter interaction. In presence of a THz electric field beyond 500 kV/cm, the generation of solvated electrons by ionization of water molecules was observed for the first time. Tunnel ionization of water molecules is induced by the strong fluctuation electric field from water dipoles reaching peak values on the order of 100 MV/cm, while the external THz field serves for a spatial transport of the released electrons away from the parent ion. The electron eventually localizes at a new site in the liquid, a process occurring on a time scale of a few picoseconds.
The solvated electron is electrically coupled to a large number of water molecules in its environment. This many-body interactions manifests in new types of collective excitations of the ensemble. Femtosecond optical pump - THz probe experiments provide the first evidence for polarons in water which are hybrid excitations of the electron and librational motions of water molecules. The polaron displays coherent oscillations in a picosecond time range and with a frequency determined by the electron concentration. The surprisingly weak damping of polaron oscillations originates from their longitudinal character and the weak coupling to other excitations of the liquid at such low frequency.
The water geometries at the surface of double-stranded helical RNA and transfer RNA (tRNA) were studied by 2D-IR spectroscopy of vibrations of the RNA backbone which serve as noninvasive probes of local interactions. The 2D-IR spectra of the asymmetric phosphate stretching vibration of tRNA reveal three prototypical hydration motifs (see Figure), a local hydration shell consisting of six water molecules forming hydrogen bonds with the free phosphate oxygens, an ordered water geometry in which individual water molecules bridge neighboring phosphate groups, and hydrated contact pairs of phosphate groups with counterions. The spectroscopic observations were analyzed by in-depth theoretical calculations and molecular dynamics simulations. RNA melting leads to a decrease of ordered structures in favor of local hydration shells. Librational water motions lead to structural fluctuations on a time scale of 300 fs while hydrogen bond lifetimes and reordering times of ions are much longer.
The work on RNA was complemented by studies of phosphate/magnesium interactions with the help of the model system dimethylphosphate. Linear and 2D-IR spectroscopy allowed for establishing clear spectroscopic signatures of contact ion pairs and underlined the particularly strong electrostatic interaction of the ions embedded in a comparably rigid water geometry. This behavior is in contrast to ion pairs with sodium and calcium ions. Currently, this work is being extended to transfer RNA structures embedded in an atmosphere of magnesium ions.
The results on the nonlinear THz response of liquid water and on the hydration structures of RNA are new and far beyond the state of the art. The water results give most direct independent insight in the role of fluctuating electric fields in the liquid and pave the way for steering electrons in water with the help of strong external fields. Our study of RNA hydration allows for the first dynamic distinction of coexisting hydration motifs and their change upon RNA melting. This insight also serves as a benchmark for theoretical work.
The work on polarons will be extended to other polar liquids to establish the potential general relevance of many-body couplings for the electric response of polar and/or ionic molecular ensembles. Nonlinear THz spectroscopy will be applied for studying the interaction of external fields with DNA and RNA, in order to quantitatively characterize the electric fields occurring in the native water environment. For this, the development of THz Stark shift spectroscopy of vibrations is underway. The combination of 2D-IR spectroscopy and theory work will allow for an in-depth characterization of hydration geometries and ion arrangements around larger RNA structures such as transfer RNA and others.
The work on polarons will be extended to other polar liquids to establish the potential general relevance of many-body couplings for the electric response of polar and/or ionic molecular ensembles. Nonlinear THz spectroscopy will be applied for studying the interaction of external fields with DNA and RNA, in order to quantitatively characterize the electric fields occurring in the native water environment. For this, the development of THz Stark shift spectroscopy of vibrations is underway. The combination of 2D-IR spectroscopy and theory work will allow for an in-depth characterization of hydration geometries and ion arrangements around larger RNA structures such as transfer RNA and others.