Investigations of Protein Fiber Structures and the Interactions at Their Interfaces Using Nonlinear Optical Spectroscopy
Johansson, Patrik Kjell Ake
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Protein fibers are ubiquitous in nature, either as functional components of cells and tissues, or as hallmarks of severe diseases. Examples include amyloid fibers in the brain of patients with Alzheimer’s disease, or collagen fibers in the extracellular matrix of tissues - central design targets for scaffolds in tissue engineering. Characterizing the structure and surface chemistry of protein fibers is therefore vital, but notoriously challenging due to their complexity. In this work, fundamental nonlinear optical properties of type I collagen fibers and amyloid structures from insulin and beta-lactoglobulin have been investigated and used for structural characterization. The collagen fibers were characterized with vibrational sum-frequency scattering spectroscopy, in a first demonstration of this technique for protein fiber investigations. Spectral features were shown to be dependent on the scattering angle and the signals away from the phase-matched direction were highly reproducible for samples with random fiber orientations. Furthermore, a scattering angle of 22˚ exhibits surface specificity for the fibers when probed in the amide I region, which was utilized to detect the initial effects of treatments with sodium dodecyl sulfate surfactants. Information from such studies of the fiber surface structure may complement the information gained from techniques with sensitivity for the overall structure, such as second-harmonic generation (SHG). For the amyloid structures, the intrinsic linear and nonlinear optical properties were investigated. In the amyloid state, beta-lactoglobulin develops a weak fluorescence in the visible regime, which could be identified by tracking the red-edge excitation shift (REES). The intrinsic optical properties allowed imaging of amyloid spherulites from both proteins, with high contrast especially for the nonlinear techniques, which included SHG and two-photon excitation fluorescence (TPEF). By monitoring the total signal from insulin amyloid spherulites in two separate detection channels while shifting the excitation wavelength, indications of the REES in TPEF were observed for the first time. The nonlinear optical techniques and phenomena explored in this work opens up for label-free detection and detailed structural investigations of protein fibers in aqueous and biological 3D environments.
- Bioengineering