Using XAS to monitor radiation damage in real time and post-analysis, and investigation of systematic errors of fluorescence XAS for Cu-bound amyloid-β

Abstract

X-ray absorption spectroscopy (XAS) is a promising technique for determining structural information from sensitive biological samples, but high-accuracy X-ray absorption fine structure (XAFS) requires corrections of systematic errors in experimental data. Low-temperature XAS and room-temperature X-ray absorption spectro-electrochemical (XAS-EC) measurements of N-truncated amyloid-β samples were collected and corrected for systematic effects such as dead time, detector efficiencies, monochromator glitches, self-absorption, radiation damage and noise at higher wavenumber (k). A new protocol was developed using extended X-ray absorption fine structure (EXAFS) data analysis for monitoring radiation damage in real time and post-analysis. The reliability of the structural determinations and consistency were validated using the XAS measurement experimental uncertainty. The correction of detector pixel efficiencies improved the fitting χ2 by 12%. An improvement of about 2.5% of the structural fitting was obtained after dead-time corrections. Normalization allowed the elimination of 90% of the monochromator glitches. The remaining glitches were manually removed. The dispersion of spectra due to self-absorption was corrected. Standard errors of experimental measurements were propagated from pointwise variance of the spectra after systematic corrections. Calculated uncertainties were used in structural refinements for obtaining precise and reliable values of structural parameters including atomic bond lengths and thermal parameters. This has permitted hypothesis testing.

Probing the Dynamic Landscape: From Static to Time‐Resolved X‐Ray Absorption Spectroscopy to Investigate Copper Redox Chemistry in Neurodegenerative Disorders

“…[38] Another way to change the Cu oxidation state stems from a factor often considered a problem for X-ray measurements with intense beams. X-ray irradiation can affect the oxidation state of Cu ions -typically by reducing Cu(II) to Cu(I) [39][40][41] -and therefore might modify the metal binding site structure.…”