Abstract
Experimental procedure and setup for obtaining X-ray fluorescence hologram of crystalline metalloprotein samples are described. Human hemoglobin, an α2β2 tetrameric metalloprotein containing the Fe(II) heme active-site in each chain, was chosen for this study because of its wealth of crystallographic data. A cold gas flow system was introduced to reduce X-ray radiation damage of protein crystals that are usually fragile and susceptible to damage. A χ-stage was installed to rotate the sample while avoiding intersection between the X-ray beam and the sample loop or holder, which is needed for supporting fragile protein crystals. Huge hemoglobin crystals (with a maximum size of 8 × 6 × 3 mm3) were prepared and used to keep the footprint of the incident X-ray beam smaller than the sample size during the entire course of the measurement with the incident angle of 0°-70°. Under these experimental and data acquisition conditions, we achieved the first observation of the X-ray fluorescence hologram pattern from the protein crystals with minimal radiation damage, opening up a new and potential method for investigating the stereochemistry of the metal active-sites in biomacromolecules.
Original language | English |
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Article number | 063707 |
Journal | Review of Scientific Instruments |
Volume | 87 |
Issue number | 6 |
DOIs | |
Publication status | Published - 2016 Jun 1 |
ASJC Scopus subject areas
- Instrumentation