The Blinking of Small-Angle X-ray Scattering Reveals the Degradation Process of Protein Crystals at Microsecond Timescale

• Published in: International journal of molecular sciences Vol. 24; no. 23; p. 16640
• Main Authors: Arai, Tatsuya, Mio, Kazuhiro, Onoda, Hiroki, Chavas, Leonard M G, Umena, Yasufumi, Sasaki, Yuji C
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 01-12-2023
• Subjects: Crystals; Lysozyme; Nanocrystals; protein crystal degradation; Proteins; Radiation; small-angle X-ray blinking (SAXB); time-resolved observations; X-ray diffraction; X-rays; Japan; X-ray diffraction; protein crystal degradation; time-resolved observations; small-angle X-ray blinking (SAXB)
• DOI: 10.3390/ijms242316640
• EISSN: 1422-0067
• ISSN: 1422-0067; 1661-6596

X-ray crystallography has revolutionized our understanding of biological macromolecules by elucidating their three-dimensional structures. However, the use of X-rays in this technique raises concerns about potential damage to the protein crystals, which results in a quality degradation of the diffraction data even at very low temperatures. Since such damage can occur on the micro- to millisecond timescale, a development in its real-time measurement has been expected. Here, we introduce diffracted X-ray blinking (DXB), which was originally proposed as a method to analyze the intensity fluctuations of diffraction of crystalline particles, to small-angle X-ray scattering (SAXS) of a lysozyme single-crystal. This novel technique, called the small-angle X-ray blinking (SAXB) method, analyzes the fluctuation in SAXS intensity reflecting the domain fluctuation in the protein crystal caused by the X-ray irradiation, which could be correlated with the X-ray-induced damage on the crystal. There was no change in the protein crystal's domain dynamics between the first and second X-ray exposures at 95K, each of which lasted 0.7 s. On the other hand, its dynamics at 295K increased remarkably. The SAXB method further showed a dramatic increase in domain fluctuations with an increasing dose of X-ray radiation, indicating the significance of this method.