Orientation Determination of Protein Helical Secondary Structures Using Linear and Nonlinear Vibrational Spectroscopy

Abstract

In this paper, we systematically presented the orientation determination of protein helical secondary structures using vibrational spectroscopic methods, particularly the nonlinear Sum Frequency Generation (SFG) vibrational spectroscopy, along with linear vibrational spectroscopic techniques such as infrared spectroscopy and Raman scattering. SFG amide I signals can be collected using different polarization combinations of the input laser beams and output signal beam to measure the second order nonlinear optical susceptibility components of the helical amide I modes, which are related to their molecular hyperpolarizability elements through the orientation distribution of these helices. The molecular hyperpolarizability elements of amide I modes of a helix can be calculated based on the infrared transition dipole moment and Raman polarizability tensor of the helix; these quantities are determined by using the bond additivity model to sum over the individual infrared dipole transition moments and Raman polarizability tensors, respectively, of the peptide units (or the amino acid residues). The computed overall infrared transition dipole moment and Raman polarizability tensor of a helix can be validated by experimental data using polarized infrared and polarized Raman spectroscopy on samples with well-aligned helical structures. From the deduced SFG hyperpolarizability elements and measured SFG second order nonlinear susceptibility components, orientation information regarding helical structures can be determined. Even though such orientation information can also be measured using polarized infrared or polarized Raman amide I signals, SFG has a much lower detection limit, which can be used to study the orientation of a helix when its surface coverage is much lower than a monolayer. In addition, the combination of different vibrational spectroscopic techniques, e.g., SFG and Attenuated Total Reflectance – Fourier Transform Infrared spectroscopy, provides more measured parameters for orientation determination, aiding in the deduction of more complicated orientation distributions. In this paper, we discussed two types of helices: the α-helix and 3–10 helix. However, the orientation determination method presented here is general, and thus can be applied to study other helices as well. The calculations of SFG amide I hyperpolarizability components for α-helical and 3–10 helical structures with different chain lengths have also been performed. It was found that when the helices reach a certain length, the number of peptide units in the helix should not alter the data analysis substantially. It was shown in the calculation, however, that when the helix chain is short, the SFG hyperpolarizability component ratios can vary substantially when the chain length is changed. Because 3–10 helical structures can be quite short in proteins, the orientation determination for a short 3–10 helix needs to take into account the number of peptide units in the helix.

Heterotrimeric G protein β 1 γ 2 subunits change orientation upon complex formation with G protein-coupled receptor kinase 2 (GRK2) on a model membrane

“…Because the SFG process is greatly enhanced by tuning the IR beam over the vibrational resonances of interfacial molecules, the observed peaks correspond to a vibrational spectrum of the interface, and highly surface-specific signals may be produced without the need for exogenous labels. By controlling the polarizations of the input and generated signal beams, information on interfacial molecular orientation can be obtained (15,16,(26)(27)(28). In our experiments, spectra were collected in the ssp and ppp polarization combinations of the sum frequency, visible, and infrared beams, respectively.…”

“…The hyperpolarizability tensor elements for each individual α-helical segment of known residue length were calculated according to the bond additivity model described previously (15). This model incorporates experimentally measured values of the dipole moment and polarizability tensor, and has been shown to produce good agreement with FTIR (33) and NMR (28) studies on simple α-helical peptides.…”

“…Recently, vibrational spectroscopic techniques such as Sum Frequency Generation (SFG) vibrational spectroscopy have been developed into a powerful, highly surface-specific, and in situ probe of biological molecules at interfaces (9)(10)(11)(12)(13)(14)(15)(16)(17). Compared to other techniques such as NMR, SFG experiments can be performed in more biologically relevant systems (single lipid bilayers), using less sample and without the requirement for isotope labeling.…”

Sticky water surfaces: Helix–coil transitions suppressed in a cell-penetrating peptide at the air-water interface

“…34 SFG has been successfully used to probe pro- tein and peptide secondary structures in the past. In those SFG studies α-helix, [35][36][37] β-sheet, 35, 37 3 10 -helix, 38 and protein aggregates [39][40][41] have been identified. Figure 3(a) shows a scheme of the experimental setup.…”

“…This problem can be avoided by directly comparing experimental SFG spectra to spectra calculated from structure files of MD simulation trajectories. 37,38 To this end we performed simulations of the full-length GALA sequence at the air-water interface (Fig. 5) in both the protonated and the deprotonated state.…”