Abstract
Kinetic studies of tau fibril formation in vitro most commonly employ spectroscopic probes such as thioflavinT fluorescence and laser light scattering or negative stain transmission electron microscopy. Here, I describe the use of Fourier transform infrared (FTIR) spectroscopy, ultraviolet resonance Raman (UVRR) spectroscopy, and atomic force microscopy (AFM) as complementary probes for studies of tau aggregation. The sensitivity of vibrational spectroscopic techniques (FTIR and UVRR) to secondary structure content allows for measurement of conformational changes that occur when the intrinsically disordered protein tau transforms into cross-β-core containing fibrils. AFM imaging serves as a gentle probe of structures populated over the time course of tau fibrillization. Together, these assays help further elucidate the structural and mechanistic complexity inherent in tau fibril formation.
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Acknowledgments
The protocols described here were standardized during graduate studies in the laboratory of Jayant B. Udgaonkar at the National Centre for Biological Sciences, Tata Institute of Fundamental Research (NCBS-TIFR), India. FTIR spectra and AFM images were acquired at the core facilities of NCBS-TIFR; UVRR spectra were collected in the laboratory of Mrinalini Puranik (then at NCBS-TIFR; now at IISER Pune, India). I’d like to acknowledge colleagues in the Puranik laboratory for teaching me the nuances of UVRR spectroscopy.
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Ramachandran, G. (2017). Fourier Transform Infrared (FTIR) Spectroscopy, Ultraviolet Resonance Raman (UVRR) Spectroscopy, and Atomic Force Microscopy (AFM) for Study of the Kinetics of Formation and Structural Characterization of Tau Fibrils. In: Smet-Nocca, C. (eds) Tau Protein. Methods in Molecular Biology, vol 1523. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6598-4_7
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DOI: https://doi.org/10.1007/978-1-4939-6598-4_7
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