Vaccinia virus immunomodulator A46 : a lipid and protein-binding scaffold for sequestering host TIR-domain proteins
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Vaccinia virus interferes with early events of the activation pathway of the transcriptional factor NF-kB by binding to numerous host TIR-domain containing adaptor proteins. We have previously determined the X-ray structure of the A46 C-terminal domain; however, the structure and function of the A46 N-terminal domain and its relationship to the C-terminal domain have remained unclear. Here, we biophysically characterize residues 1-83 of the N-terminal domain of A46 and present the X-ray structure at 1.55 Å. Crystallographic phases were obtained by a recently developed ab initio method entitled ARCIMBOLDO_BORGES that employs tertiary structure libraries extracted from the Protein Data Bank; data analysis revealed an all β-sheet structure. This is the first such structure solved by this method which should be applicable to any protein composed entirely of β-sheets. The A46(1-83) structure itself is a β-sandwich containing a co-purified molecule of myristic acid inside a hydrophobic pocket and represents a previously unknown lipid-binding fold. Mass spectrometry analysis confirmed the presence of long-chain fatty acids in both N-terminal and full-length A46; mutation of the hydrophobic pocket reduced the lipid content. Using a combination of high resolution X-ray structures of the N-and C-terminal domains and SAXS analysis of full-length protein A46(1-240), we present here a structural model of A46 in a tetrameric assembly. Integrating affinity measurements and structural data, we propose how A46 simultaneously interferes with several TIR-domain containing proteins to inhibit NF-κB activation and postulate that A46 employs a bipartite binding arrangement to sequester the host immune adaptors TRAM and MyD88.
Fedosyuk , S , Bezerra , G A , Radakovics , K , Smith , T K , Sammito , M , Bobik , N , Round , A , Ten Eyck , L F , Djinović-Carugo , K , Usón , I & Skern , T 2016 , ' Vaccinia virus immunomodulator A46 : a lipid and protein-binding scaffold for sequestering host TIR-domain proteins ' PLoS Pathogens , vol 12 , no. 12 , e1006079 . DOI: 10.1371/journal.ppat.1006079
© 2016 Fedosyuk et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
TS received Austrian Science Fund (FWF) grants P24038, W1221 and W1258. GAB is a member of Max F. Perutz Laboratories and the Vienna International PostDoctoral Program (VIPS). TKS is a holder of Wellcome Trust grant 097831. IU has Spanish Ministry of Economy and Competitiveness grant BIO2013-49604-EXP.
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