title: Rational design of crystal contact-free space in protein crystals for analyzing spatial distribution of motions within protein molecules.
abstract: Contacts with neighboring molecules in protein crystals inevitably restrict the internal motions of intrinsically flexible proteins. The resultant clear electron densities permit model building, as crystallographic snapshot structures. Although these still images are informative, they could provide biased pictures of the protein motions. If the mobile parts are located at a site lacking direct contacts in rationally designed crystals, then the amplitude of the movements can be experimentally analyzed. We propose a fusion protein method, to create crystal contact-free space (CCFS) in protein crystals and to place the mobile parts in the CCFS. Conventional model building fails when large amplitude motions exist. In this study, the mobile parts appear as smeared electron densities in the CCFS, by suitable processing of the X-ray diffraction data. We applied the CCFS method to a highly mobile presequence peptide bound to the mitochondrial import receptor, Tom20, and a catalytically relevant flexible segment in the oligosaccharyltransferase, AglB. These two examples demonstrated the general applicability of the CCFS method to the analysis of the spatial distribution of motions within protein molecules.
authors.affiliation: Division of Structural Biology, Medical Institute of Bioregulation, Kyushu University, Higashi-Ku, Fukuoka, 812-8582, Japan.
authors.forename: Rei
authors.initials: R
authors.lastname: Matsuoka
authors.affiliation: Division of Structural Biology, Medical Institute of Bioregulation, Kyushu University, Higashi-Ku, Fukuoka, 812-8582, Japan.|RIKEN Structural Biology Laboratory, Tsurumi, Yokohama, 230-0045, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Shimada
authors.affiliation: RIKEN Theoretical Molecular Science Laboratory and iTHES, Wako, Saitama, 351-0198, Japan.|Department of physics, Graduate School of Science and Engineering, Chuo University, Bunkyo-Ku, Tokyo, 112-8551, Japan.
authors.forename: Yasuaki
authors.initials: Y
authors.lastname: Komuro
authors.affiliation: RIKEN Theoretical Molecular Science Laboratory and iTHES, Wako, Saitama, 351-0198, Japan.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Sugita
authors.affiliation: Division of Structural Biology, Medical Institute of Bioregulation, Kyushu University, Higashi-Ku, Fukuoka, 812-8582, Japan.|Research Center for Live-Protein Dynamics, Kyushu University, Higashi-Ku, Fukuoka, 812-8582, Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Kohda
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D003460:Crystallization / Q000379:methods*; D018360:Crystallography, X-Ray / Q000379:methods; D006602:Hexosyltransferases / Q000737:chemistry* / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D026901:Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D024101:Mitochondrial Proteins / Q000737:chemistry* / Q000378:metabolism; D056004:Molecular Dynamics Simulation; D009038:Motion; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D020027:Pyrococcus furiosus / Q000737:chemistry* / Q000378:metabolism; D051381:Rats; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D024101:Mitochondrial Proteins; D010455:Peptides; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; C489998:Tomm20 protein, rat; D006602:Hexosyltransferases; C025620:dolichyl-diphosphooligosaccharide - protein glycotransferase
keywords: crystal contact effects; crystallographic method; fusion protein; mitochondrial presequence receptor Tom20; oligosaccharyltransferase AglB; protein motional analysis