Effects on sialic acid recognition of amino acid mutations in the carbohydrate-binding cleft of the rotavirus spike protein

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Title Effects on sialic acid recognition of amino acid mutations in the carbohydrate-binding cleft of the rotavirus spike protein
Author Kraschnefski, Mark; Bugarcic, Andrea; Fleming, Fiona E.; Yu, Xing; von Itzstein, Mark; Coulson, Barbara S.; Blanchard, Helen
Journal Name Glycobiology
Year Published 2009
Place of publication United States
Publisher Oxford University Press
Abstract The rotavirus spike protein VP4 mediates attachment to host cells and subsequent membrane penetration. The VP8* domain of VP4 forms the spike tips and is proposed to recognise host cell surface glycans. For sialidase-sensitive rotaviruses such as rhesus (RRV) this recognition involves terminal sialic acids. We show here that RRV VP8*64-224 protein competes with RRV infection of host cells, demonstrating its relevance to infection. In addition, we observe that the amino acids revealed by X-ray crystallography to be in direct contact with the bound sialic acid derivative methyl α-D-N-acetylneuraminide, and that are highly conserved amongst sialidase-sensitive rotaviruses, are residues that are also important in interactions with host-cell carbohydrates. Residues Arg101 and Ser190 of the RRV VP8* carbohydrate-binding site were mutated to assess their importance for binding to the sialic acid derivative and their competition with RRV infection of host cells. The crystallographic structure of the Arg101Ala mutant crystallised in the presence of sialic acid derivative was determined at 295 K to a resolution of 1.9 Å. Our multi-disciplinary study using X-ray crystallography, Saturation Transfer Difference nuclear magnetic resonance spectroscopy, isothermal titration calorimetry and competitive virus infectivity assays to investigate RRV wild-type and mutant VP8* proteins has provided the first evidence that the carbohydrate-binding cavity in RRV VP8* is used for host cell recognition, and this interaction is not only with the sialic acid portion but also other parts of the glycan structure.
Peer Reviewed Yes
Published Yes
Alternative URI http://dx.doi.org/10.1093/glycob/cwn119
Volume 19
Issue Number 3
Page from 194
Page to 200
ISSN 1460-2423
Date Accessioned 2009-09-16
Language en_AU
Research Centre Institute for Glycomics
Faculty Faculty of Science, Environment, Engineering and Technology
Subject Structural Biology (incl Macromolecular Modelling)
URI http://hdl.handle.net/10072/29456
Publication Type Journal Articles (Refereed Article)
Publication Type Code c1

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