Matthias Schmies

Watching water migration around a peptide bond.

Life is believed to have its origin in aqueous environments, and 70% of our body consists of water. The essential components of biological systems have to interact in aqueous solutions with water molecules by intermolecular forces, such as hydrogen bonds, dispersion forces, and hydrophilic/hydrophobic interactions. Proteins are one of the most important biological supramolecules and offer at the CO and NH sites of the -CONHlinkages of the peptide chain attractive hydrogen-bonding sites, in which H2O can act either as a proton donor or a proton acceptor, respectively. The solvation of a protein has a strong effect on its molecular shape, and as a consequence the fluctuations of the water network on the surface have important influence on its folding properties and catalytic function. Most fundamentally, when a protein starts its folding motion, the water network hydrogen-bonded to the protein has to rearrange and thus affects the dynamics. Therefore, up-to-date quantum chemical simulations on protein folding and its functions include water molecules explicitly. A deeper understanding of these phenomena at the molecular level requires the characterization of the dynamical processes of individual water molecules interacting with the protein. However, most experiments yield only indirect dynamical information averaged over water molecules in the first hydration layer and thus only a tentative and often controversial interpretation of the underlying mechanisms. Measurements visualizing the motion of a specific water molecule in a real biological environment are challenging, and so far no experimental data have been reported yet. Such dynamical experiments need to distinguish between each single water molecule, which can bind to numerous different binding sites of the protein and readily exchange their role with other H2O molecules in the same or higher hydration solvation layers. This inherent complexity of the hydrated protein has so far prevented measurements of the migration of individual water molecules in solution, and therefore nearly all information about such processes relies on theoretical approaches. Although quantum chemical simulations for such complex systems have substantially progressed in recent years because of rapid computer developments, their accuracy is still rather limited and experimental benchmark data for model systems are highly requested for calibration purposes. To this end, we have developed in the past decade an experimental strategy for the investigation of dynamical intermolecular processes, which typically occur on the picosecond (ps) time scale. This approach involves the generation of molecular clusters isolated in supersonic beams and the characterization of their dynamics using ps time-resolved IR spectroscopy. The fruitful combination of spectroscopy and quantum chemistry currently provides the most direct and most detailed access to intermolecular interactions. IR spectroscopy is particularly sensitive to structural motifs. In initial benchmark experiments, we developed a three-color UV-UV-IR tunable picosecond pump–probe laser spectrometer and measured the ionization-induced p!H site switching dynamics of rare gas ligands attached to phenol. 5] In this case, the position of the ligand was monitored by the structure-sensitive frequency of the phenolic OH stretching vibration. Although for a very limited number of water complexes with aromatic molecules the laser-induced migration of the water ligand has recently been inferred from “static” spectroscopy using nanosecond lasers, 6] no time-resolved studies about the dynamics of this fundamental process have been reported yet. We have applied our ps pump–probe approach to the trans-acetanilide–H2O (AA-H2O) cluster to monitor the water migration dynamics around a peptide linkage by the time evolution of the IR spectra measured at a delay time Dt after the ionization event (Figure 1). AA is one of the smallest aromatic molecules with a -CONHpeptide bond (Figure 1) and thus serves as a suitable model for proteins and related biomolecules. It can form two different types of hydrogen bonds with water, resulting in NHand CO-bound isomers, which can readily be distinguished by their different IR and electronic spectra 7] (see the Supporting Information). The “static” IR spectrum of the CO-bound isomer (reactand R) in the neutral ground state (S0) measured by nanosecond lasers (Dt = 50 ns, Figure 2) reveals a sharp band at 3473 cm 1 and a broader one at 3496 cm 1 assigned to the free NH stretching vibration (nNH ) and the hydrogen-bonded OH stretching mode (nOH ) of the water ligand, respectively. The corresponding picosecond IR spectra before photoionization [*] K. Tanabe, Dr. M. Miyazaki, Prof. Dr. M. Sakai, Prof. Dr. M. Fujii Chemical Resources Laboratory, Tokyo Institute of Technology Yokohama 226-8503 (Japan) E-mail: mfujii@res.titech.ac.jp

Structures and IR/UV spectra of neutral and ionic phenol-Ar(n) cluster isomers (n ≤ 4): competition between hydrogen bonding and stacking.

The structures, binding energies, and vibrational and electronic spectra of various isomers of neutral and ionic phenol-Ar(n) clusters with n ≤ 4, PhOH((+))-Ar(n), are characterized by quantum chemical calculations. The properties in the neutral and ionic ground electronic states (S(0), D(0)) are determined at the M06-2X/aug-cc-pVTZ level, whereas the S(1) excited state of the neutral species is investigated at the CC2/aug-cc-pVDZ level. The Ar complexation shifts calculated for the S(1) origin and the adiabatic ionisation potential, ΔS(1) and ΔIP, sensitively depend on the Ar positions and thus the sequence of filling the first Ar solvation shell. The calculated shifts confirm empirical additivity rules for ΔS(1) established recently from experimental spectra and enable thus a firm assignment of various S(1) origins to their respective isomers. A similar additivity model is newly developed for ΔIP using the M06-2X data. The isomer assignment is further confirmed by Franck-Condon simulations of the intermolecular vibrational structure of the S(1) ← S(0) transitions. In neutral PhOH-Ar(n), dispersion dominates the attraction and π-bonding is more stable than H-bonding. The solvation sequence of the most stable isomers is derived as (10), (11), (30), and (31) for n ≤ 4, where (km) denotes isomers with k and m Ar ligands binding above and below the aromatic plane, respectively. The π interaction is somewhat stronger in the S(1) state due to enhanced dispersion forces. Similarly, the H-bond strength increases in S(1) due to the enhanced acidity of the OH proton. In the PhOH(+)-Ar(n) cations, H-bonds are significantly stronger than π-bonds due to additional induction forces. Consequently, one favourable solvation sequence is derived as (H00), (H10), (H20), and (H30) for n ≤ 4, where (Hkm) denotes isomers with one H-bound ligand and k and m π-bonded Ar ligands above and below the aromatic plane, respectively. Another low-energy solvation motif for n = 2 is denoted (11)(H) and involves nonlinear bifurcated H-bonding to both equivalent Ar atoms in a C(2v) structure in which the OH group points toward the midpoint of an Ar(2) dimer in a T-shaped fashion. This dimer core can also be further solvated by π-bonded ligands leading to the solvation sequence (H00), (11)(H), (21)(H), and (22) for n ≤ 4. The implications of the ionisation-induced π → H switch in the preferred interaction motif on the isomerisation and fragmentation processes of PhOH((+))-Ar(n) are discussed in the light of the new structural and energetic cluster parameters.

Microhydrated aromatic cluster cations: Binding motifs of 4-aminobenzonitrile-(H2O)n cluster cations with n ≤ 4

Infrared photodissociation (IRPD) spectra of mass-selected 4-aminobenzonitrile-(water)n cluster cations, ABN(+)-(H2O)n with n ≤ 4, recorded in the N-H and O-H stretch ranges are analyzed by quantum chemical calculations at the M06-2X/aug-cc-pVTZ level to determine the evolution of the initial microhydration process of this bifunctional aromatic cation in its ground electronic state. IRPD spectra of cold clusters tagged with Ar and N2 display higher resolution and allow for a clear-cut structural assignment. The clusters are generated in an electron impact source, which generates predominantly the most stable isomers. The IRPD spectra are assigned to single isomers for n = 1-3. The preferred cluster growth begins with sequential hydration of the two acidic NH protons of the amino group (n = 1-2), which is followed by attachment of secondary H2O ligands hydrogen-bonded to the first-shell ligands (n = 3-4). These symmetric and branched structures are more stable than those with a cyclic H-bonded solvent network. Moreover, in the size range n ≤ 4 the formation of a solvent network stabilized by strong cooperative effects is favored over interior ion hydration which is destabilized by noncooperative effects. The potential of the ABN(+)-H2O dimer is characterized in detail and supports the cluster growth derived from the IRPD spectra. Although the N-H bonds are destabilized by stepwise microhydration, which is accompanied by increasing charge transfer from ABN(+) to the solvent cluster, no proton transfer to the solvent is observed for n ≤ 4.

Stepwise Microhydration of Aromatic Amide Cations: Formation of Water Solvation Network Revealed by Infrared Spectra of Formanilide+–(H2O)n Clusters (n ≤ 5)

Hydration of peptides and proteins has a strong impact on their structure and function. Infrared photodissociation spectra (IRPD) of size-selected clusters of the formanilide cation, FA(+)-(H2O)n (n = 1-5), are analyzed by density functional theory calculations at the ωB97X-D/aug-cc-pVTZ level to determine the sequential microhydration of this prototypical aromatic amide cation. IRPD spectra are recorded in the hydride stretch and fingerprint ranges to probe the preferred interaction motifs and the cluster growth. IRPD spectra of cold Ar-tagged clusters, FA(+)-(H2O)n-Ar, reveal the important effects of temperature and entropy on the observed hydration motifs. At low temperature, the energetically most stable isomers are prominent, while at higher temperature less stable but more flexible isomers become increasingly populated because of entropy. In the most stable structures, the H2O ligands form a hydrogen-bonded solvent network attached to the acidic NH proton of the amide, which is stabilized by large cooperative effects arising from the excess positive charge. In larger clusters, hydration bridges the gap between the NH and CO groups (n ≥ 4) solvating the amide group rather than the more positively charged phenyl ring. Comparison with neutral FA-(H2O)n clusters reveals the strong impact of ionization on the acidity of the NH proton, the strength and topology of the interaction potential, and the structure of the hydration shell.

Weak hydrogen bonding motifs of ethylamino neurotransmitter radical cations in a hydrophobic environment: infrared spectra of tryptamine(+)-(N2)n clusters (n ≤ 6).

Size-selected clusters of the tryptamine cation with N2 ligands, TRA(+)-(N2)n with n = 1-6, are investigated by infrared photodissociation (IRPD) spectroscopy in the hydride stretch range and quantum chemical calculations at the ωB97X-D/cc-pVTZ level to characterize the microsolvation of this prototypical aromatic ethylamino neurotransmitter radical cation in a nonpolar solvent. Two types of structural isomers exhibiting different interaction motifs are identified for the TRA(+)-N2 dimer, namely the TRA(+)-N2(H) global minimum, in which N2 forms a linear hydrogen bond (H-bond) to the indolic NH group, and the less stable TRA(+)-N2(π) local minima, in which N2 binds to the aromatic π electron system of the indolic pyrrole ring. The IRPD spectrum of TRA(+)-(N2)2 is consistent with contributions from two structural H-bound isomers with similar calculated stabilization energies. The first isomer, denoted as TRA(+)-(N2)2(2H), exhibits an asymmetric bifurcated planar H-bonding motif, in which both N2 ligands are attached to the indolic NH group in the aromatic plane via H-bonding and charge-quadrupole interactions. The second isomer, denoted as TRA(+)-(N2)2(H/π), has a single and nearly linear H-bond of the first N2 ligand to the indolic NH group, whereas the second ligand is π-bonded to the pyrrole ring. The natural bond orbital analysis of TRA(+)-(N2)2 reveals that the total stability of these types of clusters is not only controlled by the local H-bond strengths between the indolic NH group and the N2 ligands but also by a subtle balance between various contributing intermolecular interactions, including local H-bonds, charge-quadrupole and induction interactions, dispersion, and exchange repulsion. The systematic spectral shifts as a function of cluster size suggest that the larger TRA(+)-(N2)n clusters with n = 3-6 are composed of the strongly bound TRA(+)-(N2)2(2H) core ion to which further N2 ligands are weakly attached to either the π electron system or the indolic NH proton by stacking and charge-quadrupole forces.

Simultaneous Interaction of Hydrophilic and Hydrophobic Solvents with Ethylamino Neurotransmitter Radical Cations: Infrared Spectra of Tryptamine(+)-(H2O)m-(N2)n Clusters (m,n ≤ 3).

Solvation of biomolecules by a hydrophilic and hydrophobic environment strongly affects their structure and function. Here, the structural, vibrational, and energetic properties of size-selected clusters of the microhydrated tryptamine cation with N2 ligands, TRA(+)-(H2O)m-(N2)n (m,n ≤ 3), are characterized by infrared photodissociation spectroscopy in the 2800-3800 cm(-1) range and dispersion-corrected density functional theory calculations at the ωB97X-D/cc-pVTZ level to investigate the simultaneous solvation of this prototypical neurotransmitter by dipolar water and quadrupolar N2 ligands. In the global minimum structure of TRA(+)-H2O generated by electron ionization, H2O is strongly hydrogen-bonded (H-bonded) as proton acceptor to the acidic indolic NH group. In the TRA(+)-H2O-(N2)n clusters, the weakly bonded N2 ligands do not affect the H-bonding motif of TRA(+)-H2O and are preferentially H-bonded to the OH groups of the H2O ligand, whereas stacking to the aromatic π electron system of the pyrrole ring of TRA(+) is less favorable. The natural bond orbital analysis reveals that the H-bond between the N2 ligand and the OH group of H2O cooperatively strengthens the adjacent H-bond between the indolic NH group of TRA(+) and H2O, while π stacking is slightly noncooperative. In the larger TRA(+)-(H2O)m clusters, the H2O ligands form a H-bonded solvent network attached to the indolic NH proton, again stabilized by strong cooperative effects arising from the nearby positive charge. Comparison with the corresponding neutral TRA-(H2O)m clusters illustrates the strong impact of the excess positive charge on the structure of the microhydration network.

Solvent migration in microhydrated aromatic aggregates: ionization-induced site switching in the 4-aminobenzonitrile-water cluster.

The dependence of the preferred microhydration sites of 4-aminobenzonitrile (4ABN) on electronic excitation and ionization is determined through IR spectroscopy of its clusters with water (W) in a supersonic expansion and through quantum chemical calculations. IR spectra of neutral 4ABN and two isomers of its hydrogen-bonded (H-bonded) 4ABN-W complexes are obtained in the ground and first excited singlet states (S0, S1) through IR depletion spectroscopy associated with resonance-enhanced multiphoton ionization. Spectral analysis reveals that electronic excitation does not change the H-bonding motif of each isomer, that is, H2O binding either to the CN or the NH site of 4ABN, denoted as 4ABN-W(CN) and 4ABN-W(NH), respectively. The IR spectra of 4ABN(+)-W in the doublet cation ground electronic state (D0) are measured by generating them either in an electron ionization source (EI-IR) or through resonant multiphoton ionization (REMPI-IR). The EI-IR spectrum shows only transitions of the most stable isomer of the cation, which is assigned to 4ABN(+)-W(NH). The REMPI-IR spectrum obtained through isomer-selective resonant photoionization of 4ABN-W(NH) is essentially the same as the EI-IR spectrum. The REMPI-IR spectrum obtained by ionizing 4ABN-W(CN) is also similar to that of the 4ABN(+)-W(NH) isomer, but differs from that calculated for 4ABN(+)-W(CN), indicating that the H2O ligand migrates from the CN to the NH site upon ionization with a yield of 100%. The mechanism of this CN→NH site-switching reaction is discussed in the light of the calculated potential energy surface and the role of intracluster vibrational energy redistribution.

Microsolvation of the Formanilide Cation (FA+) in a Nonpolar Solvent: Infrared Spectra of FA+–Ln Clusters (L = Ar, N2; n ≤ 8)

Infrared photodissociation (IRPD) spectra of cationic formanilide (N-phenylformamide) clusters, FA(+)-Ln, with L = Ar (n = 1-8) and N2 (n = 1-6), are recorded in the hydride stretch (amide A, νNH, νCH) and fingerprint (amide I-III) ranges to probe the preferred interaction motifs and the cluster growth. Cold FA(+)-Ln clusters are generated by electron ionization in a supersonic expansion, which generates predominantly the most stable cluster isomers. Size- and isomer-specific νNH frequencies unravel the microsolvation process of FA(+) in a nonpolar (L = Ar) and a quadrupolar (L = N2) solvent. The H-bound FA(+)-L dimer with L binding to the NH proton of the amide group is the most stable isomer, and further ligands are attached to the aromatic ring (π-stacking). Ionization changes the preferred binding motif from π-stacking to H-bonding in FA((+))-L. Quantum chemical calculations at the ωB97X-D/aug-cc-pVTZ level confirm the experimentally derived sequential cluster growth and the vibrational and isomer assignments. The calculated FA(+)-L binding energies of D0(H) = 594/1054 cm(-1) for H-bound and D0(π) = 459/604 cm(-1) for π-bound Ar/N2 ligands are consistent with the observed photofragmentation branching ratios. Ionization of FA results from removal of a bonding π-electron delocalized over the phenyl and amide moieties and thus weakens the N-H bond and strengthens the C-O bond.

Microsolvation of the 4‐Aminobenzonitrile Cation (ABN+) in a Nonpolar Solvent: IR Spectra of ABN+Ln (L=Ar and N2, n≤4)

IR photodissociation (IRPD) spectra of mass-selected cluster ions of 4-aminobenzonitrile (ABN(+)) with up to four Ar and N2 ligands are recorded over the spectral range of the N-H stretching vibrations (ν(s/a)) of ABN(+) in its (2)B1 ground electronic state. ABN(+)-L(n) clusters are produced in an electron impact cluster ion source, which predominantly generates the most stable isomer of a given cluster ion. Vibrational frequency shifts of ν(s/a) provide information about the sequential microsolvation process of ABN(+) in a nonpolar solvent. In ABN(+)-(N2)n, the first two ligands fill a first subshell by forming hydrogen bonds to the acidic protons of the amino group, whereas further ligands bind more weakly to the aromatic ring (π bonds). Although the preferred cluster growth sequence in ABN(+)-Ar(n) is similar, several isomers are observed because the hydrogen bonds are only slightly stronger than the π bonds. Quantum chemical calculations at the M06-2X/aug-cc-pVTZ level confirm the cluster growth sequence derived from the IR spectra and provide further details of the intermolecular potential. The calculated binding energies of D0(H)=532 and 895 cm(-1) for hydrogen-bonded and D0(π)=512 and 530 cm(-1) for π-bonded Ar and N2 ligands are consistent with the observed photofragmentation branching ratios. Comparison between ABN(+)-L(n) and the corresponding clusters with the aniline cation demonstrates that the NH protons of the amino group become slightly more acidic upon H→CN substitution at the para position. Comparison between charged and neutral ABN((+))-L dimers indicates that ionization switches the preferred ion-ligand binding motif from π to hydrogen bonding.

IR spectroscopy of the 4-aminobenzonitrile-Ar cluster in the S0, S1 neutral and D0 cationic states.

The S1-S0 resonant enhanced multiphoton ionization (REMPI) spectrum as well as the infrared (IR) spectra in the S0 and S1 states of 4-aminobenzonitrile (4ABN) and its van der Waals complex with Ar (4ABN-Ar) were measured by means of IR depletion spectroscopy (REMPI-IR). The IR spectrum of 4ABN-Ar in S0 shows symmetric and antisymmetric NH stretching vibrations (ν(s) and ν(a)) of the amino group at the same positions as those in the 4ABN monomer. This suggests that the Ar ligand locates above the benzene ring by van der Waals interactions (π-bound). The same coincidence of vibrational frequencies was found in S1, and the π-bound geometry was kept by the electronic excitation. The REMPI-IR spectrum of 4ABN(+)-Ar was also measured, and three major vibrational transitions were found. From the comparison to the IR dissociation spectrum with an electron impact source (EI-IR), they were assigned to ν(s), ν(a) and an NH-bending overtone of the π-bound structure. It is concluded that photoionization of 4ABN(+)-Ar does not promote site-switching of Ar from the π-site to the H-site.