title: Development of an X-ray fluorescence holographic measurement system for protein crystals.
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 mm(3)) 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.
authors.affiliation: Division of Biophysics, Department of Physiology, Jichi Medical University, Yakushiji, Shimotsuke 329-0498, Japan.
authors.forename: Ayana
authors.initials: A
authors.lastname: Sato-Tomita
authors.affiliation: Division of Biophysics, Department of Physiology, Jichi Medical University, Yakushiji, Shimotsuke 329-0498, Japan.
authors.forename: Naoya
authors.initials: N
authors.lastname: Shibayama
authors.affiliation: Department of Computer and Network Engineering, Graduate School of Information Sciences, Hiroshima City University, Asa-Minami-Ku, Hiroshima 731-3194, Japan.
authors.forename: Naohisa
authors.initials: N
authors.lastname: Happo
authors.affiliation: Department of Physical Science and Engineering, Nagoya Institute of Technology, Gokiso, Showa, Nagoya 466-8555, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Kimura
authors.affiliation: Division of Biophysics, Department of Physiology, Jichi Medical University, Yakushiji, Shimotsuke 329-0498, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Okabe
authors.affiliation: Japan Synchrotron Radiation Research Institute (JASRI), SPring-8, Sayo, Hyogo 679-5198, Japan.
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Matsushita
authors.affiliation: Drug Design Laboratory, Department of Medical Life Science, Yokohama City University, Suehiro, Tsurumi, Yokohama 230-0045, Japan.
authors.forename: Sam-Yong
authors.initials: SY
authors.lastname: Park
authors.affiliation: Department of Advanced Material Science, Graduate School of Frontier Science, The University of Tokyo, Kashiwanoha, Kashiwa 277-8561, Japan.
authors.forename: Yuji C
authors.initials: YC
authors.lastname: Sasaki
authors.affiliation: Department of Physical Science and Engineering, Nagoya Institute of Technology, Gokiso, Showa, Nagoya 466-8555, Japan.
authors.forename: Kouichi
authors.initials: K
authors.lastname: Hayashi
mesh_terms: D020134:Catalytic Domain; D006454:Hemoglobins / Q000737:chemistry*; D006696:Holography / Q000379:methods*; D006801:Humans; D013052:Spectrometry, X-Ray Emission / Q000379:methods*
chemical_list: D006454:Hemoglobins