Abstract
Measles virus (MV), one of the most contagious agents, infects immune cells using the signaling lymphocyte activation molecule (SLAM) on the cell surface. A complex of SLAM and the attachment protein, hemagglutinin (MVH), has remained elusive due to the intrinsic handling difficulty including glycosylation. Furthermore, crystals obtained of this complex are either nondiffracting or poorly-diffracting. To solve this problem, we designed a systematic approach using a combination of the following techniques; (1) a transient expression system in HEK293SGnTI(-) cells, (2) lysine methylation, (3) structure-guided mutagenesis directed at better crystal packing, (4) Endo H treatment, (5) single-chain formation for stable complex, and (6) floating-drop vapor diffusion. Using our approach, the receptor-binding head domain of MV-H covalently fused with SLAM was successfully crystallized and diffraction was improved from 4.5 Å to a final resolution of 3.15 Å . These combinational methods would be useful as crystallization strategies for complexes of glycoproteins and their receptors.
Keywords: SLAM, Crystallization, floating-drop vapor diffusion, glycoprotein, HEK293SGnTI(-) cells, lysine methylation, measles, virus, hemagglutinin, receptor
Protein & Peptide Letters
Title:Crystallization Strategy for the Glycoprotein-Receptor Complex Between Measles Virus Hemagglutinin and Its Cellular Receptor SLAM
Volume: 19 Issue: 4
Author(s): Takao Hashiguchi, Toyoyuki Ose, Marie Kubota, Nobuo Maita, Jun Kamishikiryo, Katsumi Maenaka and Yusuke Yanagi
Affiliation:
Keywords: SLAM, Crystallization, floating-drop vapor diffusion, glycoprotein, HEK293SGnTI(-) cells, lysine methylation, measles, virus, hemagglutinin, receptor
Abstract: Measles virus (MV), one of the most contagious agents, infects immune cells using the signaling lymphocyte activation molecule (SLAM) on the cell surface. A complex of SLAM and the attachment protein, hemagglutinin (MVH), has remained elusive due to the intrinsic handling difficulty including glycosylation. Furthermore, crystals obtained of this complex are either nondiffracting or poorly-diffracting. To solve this problem, we designed a systematic approach using a combination of the following techniques; (1) a transient expression system in HEK293SGnTI(-) cells, (2) lysine methylation, (3) structure-guided mutagenesis directed at better crystal packing, (4) Endo H treatment, (5) single-chain formation for stable complex, and (6) floating-drop vapor diffusion. Using our approach, the receptor-binding head domain of MV-H covalently fused with SLAM was successfully crystallized and diffraction was improved from 4.5 Å to a final resolution of 3.15 Å . These combinational methods would be useful as crystallization strategies for complexes of glycoproteins and their receptors.
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Hashiguchi Takao, Ose Toyoyuki, Kubota Marie, Maita Nobuo, Kamishikiryo Jun, Maenaka Katsumi and Yanagi Yusuke, Crystallization Strategy for the Glycoprotein-Receptor Complex Between Measles Virus Hemagglutinin and Its Cellular Receptor SLAM, Protein & Peptide Letters 2012; 19 (4) . https://dx.doi.org/10.2174/092986612799789314
DOI https://dx.doi.org/10.2174/092986612799789314 |
Print ISSN 0929-8665 |
Publisher Name Bentham Science Publisher |
Online ISSN 1875-5305 |
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