Patrick S. Walsh

Role of ring-constrained γ-amino acid residues in α/γ-peptide folding: single-conformation UV and IR spectroscopy.

The capped α/γ-peptide foldamers Ac-γACHC-Ala-NH-benzyl (γα) and Ac-Ala-γACHC-NH-benzyl (αγ) were studied in the gas phase under jet-cooled conditions using single-conformation spectroscopy. These molecules serve as models for local segments of larger heterogeneous 1:1 α/γ-peptides that have recently been synthesized and shown to form a 12-helix composed of repeating C12 H-bonded rings both in crystalline form and in solution [Guo, L.; et al. J. Am. Chem. Soc. 2009, 131, 16018]. The γα and αγ peptide subunits are structurally constrained at the Cβ-Cγ bond of the γ-residue with a cis-cyclohexyl ring and by an ethyl group at the Cα position. These triamides are the minimum length necessary for the formation of the C12 H-bond. Resonant two-photon ionization (R2PI) provides ultraviolet spectra that have contributions from all conformational isomers, while IR-UV hole-burning (IR-UV HB) and resonant ion-dip infrared (RIDIR) spectroscopies are used to record single-conformation UV and IR spectra, respectively. Four and six conformers are identified in the R2PI spectra of the γα and αγ peptides, respectively. RIDIR spectra in the NH stretch, amide I (C═O stretch), and amide II (NH bend) regions are compared with the predictions of density functional theory (DFT) calculations at the M05-2X/6-31+G* level, leading to definite assignments for the H-bonding architectures of the conformers. While the C12 H-bond is present in both γα and αγ, C9 rings are more prevalent, with seven of ten conformers incorporating a C9 H-bond involving in the γ-residue. Nevertheless, comparison of the assigned structures of gas-phase γα and αγ with the crystal structures for γα and larger α/γ-peptides reveals that the constrained γ-peptide backbone formed by the C9 ring is structurally similar to that formed by the larger C12 ring present in the 12-helix. These results confirm that the ACHC/ethyl constrained γ-residue is structurally preorganized to play a significant role in promoting C12 H-bond formation in larger α/γ-peptides.

Cyclic Constraints on Conformational Flexibility in γ-Peptides: Conformation Specific IR and UV Spectroscopy

Single-conformation spectroscopy has been used to study two cyclically constrained and capped γ-peptides: Ac-γACHC-NHBn (hereafter γACHC, Figure 1a), and Ac-γACHC-γACHC-NHBn (γγACHC, Figure 1b), under jet-cooled conditions in the gas phase. The γ-peptide backbone in both molecules contains a cyclohexane ring incorporated across each Cβ-Cγ bond and an ethyl group at each Cα. This substitution pattern was designed to stabilize a (g+, g+) torsion angle sequence across the Cα-Cβ-Cγ segment of each γ-amino acid residue. Resonant two-photon ionization (R2PI), infrared-ultraviolet hole-burning (IR-UV HB), and resonant ion-dip infrared (RIDIR) spectroscopy have been used to probe the single-conformation spectroscopy of these molecules. In both γACHC and γγACHC, all population is funneled into a single conformation. With RIDIR spectra in the NH stretch (3200-3500 cm(-1)) and amide I/II regions (1400-1800 cm(-1)), in conjunction with theoretical predictions, assignments have been made for the conformations observed in the molecular beam. γACHC forms a single nearest-neighbor C9 hydrogen-bonded ring whereas γγACHC takes up a next-nearest-neighbor C14 hydrogen-bonded structure. The gas-phase C14 conformation represents the beginning of a 2.614-helix, suggesting that the constraints imposed on the γ-peptide backbone by the ACHC and ethyl groups already impose this preference in the gas-phase di-γ-peptide, in which only a single C14 H-bond is possible, constituting one full turn of the helix. A similar conformational preference was previously documented in crystal structures and NMR analysis of longer γ-peptide oligomers containing the γACHC subunit [Guo, L., et al. Angew. Chem. Int. Ed. 2011, 50, 5843-5846]. In the gas phase, the γACHC-H2O complex was also observed and spectroscopically interrogated in the molecular beam. Here, the monosolvated γACHC retains the C9 hydrogen bond observed in the bare molecule, with the water acting as a bridge between the C-terminal carbonyl and the π-cloud of the UV chromophore. This is in contrast to the unconstrained γ-peptide-H2O complex, which incorporates H2O into both C9 and amide-stacked conformations.

Mimicking the First Turn of an α-Helix with an Unnatural Backbone: Conformation-Specific IR and UV Spectroscopy of Cyclically Constrained β/γ-Peptides

The folding preferences of two capped, constrained β/γ-dipeptide isomers, Ac-βACPC-γACHC-NHBn and Ac-γACHC-βACPC-NHBn, (designated βγ and γβ, respectively), have been investigated using single- and double-resonance ultraviolet and infrared spectroscopy in the gas phase. These capped β/γ-dipeptides have the same number of backbone atoms between their N- and C-termini as a capped α-tripeptide and thus serve as a minimal structural unit on which to test their ability to mimic the formation of the first turn of an α-helix. Resonant two-photon ionization and UV-UV hole-burning spectroscopy were performed in the S0-S1 region, revealing the presence of three unique conformations of βγ and a single conformation of γβ. Resonant ion-dip infrared spectra were obtained in the NH stretch region from 3300 to 3500 cm(-1) and in both the amide I and amide II regions from 1400 to 1800 cm(-1). These infrared spectra were compared to computational predictions from density functional theory calculations at the M05-2X/6-31+G(d) level, leading to assignments for the observed conformations. Two unique bifurcated C8/C13 H-bonded ring structures for βγ and a single bifurcated C9/C13 H-bonded ring structure for γβ were observed. In all cases, the H-bonding patterns faithfully mimic the first full turn of an α-helix, most notably by containing a 13-membered H-bonded cycle but also by orienting the interior amide group so that it is poised to engage in a second C13 H-bond as the β/γ-peptide lengthens in size. The structural characteristics of the β/γ-peptide version of the 13-helix turn are compared with the α-helix counterpart and with a reported crystal structure for a longer β/γ-peptide oligomer.

Local Mode Approach to OH Stretch Spectra of Benzene-(H2O)n Clusters, n = 2-7.

Isomer-specific resonant ion-dip infrared spectra are presented for benzene-water (Bz-(H2O)n) clusters with two to seven water molecules. Local mode Hamiltonians based on scaled M06-2X/6-311++G(2d,p) density functional calculations are presented that accurately model the spectra across the entire OH stretch region (3000-3750 cm(-1)). The model Hamiltonians include the contribution from the water bend overtone and an empirical parameter for the local OH stretch-bend Fermi coupling. The inclusion of this coupling is necessary for accurate modeling of the infrared spectra of clusters with more than three water molecules. For the cyclic water clusters (n = 3-5), the benzene molecule perturbs the system in a characteristic way, distorting the cycle, splitting degeneracies, and turning on previously forbidden transitions. The local OH stretch site frequencies and H···OH hydrogen bond lengths follow a pattern based on the each water monomers proximity to benzene. The patterns observed for these cyclic water clusters provide insight into benzenes effects on the three-dimensional hydrogen-bonded networks present in water hexamer and heptamer structures, which also have their spectra dramatically altered from their pure water counterparts.

Single-conformation UV and IR spectroscopy of model G-type lignin dilignols: the β–O–4 and β–β linkages

Single-conformation ultraviolet and infrared spectroscopy was performed on dilignols containing two of the three biologically prevalent β-lignol linkages, erythro β–O–4 (β-aryl ether) and (±) β–β (pinoresinol). Both dilignols contain guaiacol(G)-type sub-units, representative of these linkages in G-type lignin. Resonant two-photon ionization (R2PI), IR-UV, and UV-UV holeburning (UVHB) spectroscopy in the cold, isolated environment of a supersonic expansion was carried out to determine the spectroscopic signatures associated with each linkage conformation, revealing striking differences in the vibronic intensity patterns between the two molecules in the UV. Two conformational isomers were found for the β–O–4 dilignol, both being classified into the fully hydrogen-bonded family with α-OH⋯OCH3 (C8) and γ-OH⋯Oβ (C5) H-bonds that are characteristic of the β–O–4 linkage. Conversely, a single dominant conformation was found for the conformationally-constrained pinoresinol. Resonant ion-dip infrared (RIDIR) spectroscopy provided conformation-specific IR spectra in the OH stretch and alkyl CH stretch regions, yielding complementary data that reported on both the intramolecular H-bonding and more subtle linkage features, respectively. DFT M05-2X calculations predict that the rigid β–β linkage had far fewer low-energy conformations in the first 20 kJ mol−1 (3) than the more flexible β–O–4 linkage (45). In the β–O–4 lignin dimer, the distinct UV chromophores lead to a splitting between S1 and S2 states that is determined mainly by the differences in the chemical structures of the two chromophores. In pinoresinol however, the assigned structure has C2 symmetry, with a calculated vertical excitonic splitting between the S1 and S2 states of 74 cm−1 (TDDFT). After taking into account the reduction in splitting associated with the geometry change in the aromatic rings upon electronic excitation, a vibronically quenched excitonic splitting of no more than a few wavenumbers is predicted for the C2 symmetric pinoresinol, but a definite experimental confirmation was not possible. These results predict that, under most circumstances, adjacent chromophores along a lignin polymer chain are not significantly electronically coupled to one another, and can be treated largely as isolated chromophores.

Inherent Conformational Preferences of Ac-Gln-Gln-NHBn: Sidechain Hydrogen Bonding Supports a β-Turn in the Gas Phase

Gas-phase single-conformation spectroscopy is used to study Ac-Gln-Gln-NHBn in order to probe the interplay between sidechain hydrogen bonding and backbone conformational preferences. This small, amide-rich peptide offers many possibilities for backbone-backbone, sidechain-backbone, and sidechain-sidechain interactions. The major conformer observed experimentally features a type-I β-turn with a canonical 10-membered ring C=O-H-N hydrogen bond between backbone amide groups. In addition, the C=O group of each Gln sidechain participates in a seven-membered ring hydrogen bond with the backbone NH of the same residue. Thus, sidechain hydrogen-bonding potential is satisfied in a manner that is consistent with and stabilizes the β-turn secondary structure. This turn-forming propensity may be relevant to pathogenic amyloid formation by polyglutamine segments in human proteins.

Conformer-Specific and Diastereomer-Specific Spectroscopy of αβα Synthetic Foldamers: Ac-Ala-βACHC-Ala-NHBn

The folding propensities of a capped, cyclically constrained, mixed α/β diastereomer pair, ( SRSS) Ac-Ala-βACHC-Ala-NHBn (hereafter RS) and ( SSRS) Ac-Ala-βACHC-Ala-NHBn ( SR), have been studied in a molecular beam using single-conformation spectroscopic techniques. These α/β-tripeptides contain a cyclohexane ring across each Cα -Cβ bond, at which positions their stereochemistries differ. This cyclic constraint requires any stable species to adopt one of two ACHC configurations: equatorial C═O/axial NH or equatorial NH/axial C═O. Resonant two-photon ionization (R2PI) and infrared-ultraviolet hole-burning (IR-UV HB) spectroscopy were used in the S0-S1 region of the UV chromophore, revealing the presence of three unique conformational isomers of RS and two of SR. Resonant ion-dip infrared spectra were recorded in both the NH stretch (3200-3500 cm-1) and the amide I (1600-1800 cm-1) regions. These experimental vibrational frequencies were compared with the scaled calculated normal-mode frequencies from density functional theory at the M05-2X/6-31+G(d) level of theory, leading to structural assignments of the observed conformations. The RS diastereomer is known in crystalline form to preferentially form a C11/C9 mixed helix, in which alternating hydrogen bonds are arranged in near antiparallel orientation. This structure is preserved in one of the main conformers observed in the gas phase but is in competition with both a tightly folded C7eq/C12/C8/C7eq structure, in which all four amide NH groups and four C═O groups are engaged in hydrogen bonding, as well as a cap influenced C7eq/NH···π/C11 structure. The SR diastereomer is destabilized by inducing backbone dihedral angles that lie outside the typical Ramachandran angles. This diastereomer also forms a C11/C9 mixed helix as well as a cap influenced bifurcated C7ax-C11/NH···π/C7eq structure as the global energy minimum. Assigned structures are compared with the reported crystal structure of analogous α/β-tripeptides, and disconnectivity graphs are presented to give an overview of the complicated potential energy surface of this tripeptide diastereomer pair.