SASDBK7: Vaccinia virus A46 protein (full-length) (Protein A46, VACV A46)

Basic information

• Entry: Database: SASBDB / ID: SASDBK7

Sample

Vaccinia virus A46 protein (full-length)

• Protein A46 (protein), VACV A46, Vaccinia virus

• Function / homology: Poxvirus Bcl-2-like / Poxvirus domain superfamily / Poxvirus Bcl-2-like proteins / extrinsic component of cytoplasmic side of plasma membrane / protein sequestering activity / symbiont-mediated suppression of host toll-like receptor signaling pathway / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of IRF3 activity / virus-mediated perturbation of host defense response / Protein OPG176
• Biological species: Vaccinia virus
• Citation: Journal: PLoS Pathog / Year: 2016; Title: Vaccinia Virus Immunomodulator A46: A Lipid and Protein-Binding Scaffold for Sequestering Host TIR-Domain Proteins.; Authors: Sofiya Fedosyuk / Gustavo Arruda Bezerra / Katharina Radakovics / Terry K Smith / Massimo Sammito / Nina Bobik / Adam Round / Lynn F Ten Eyck / Kristina Djinović-Carugo / Isabel Usón / Tim Skern / ; Abstract: 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.

Contact author

• Sofiya Fedosyuk (Medical University of Vienna, Vienna, Austria)

Models

• Model #854: ; Type: mix / Software: CORAL / Radius of dummy atoms: 1.90 A / Chi-square value: 0.54; Search similar-shape structures of this assembly by Omokage search (details)
• Model #853: ; Type: mix / Software: CORAL / Radius of dummy atoms: 1.90 A / Chi-square value: 0.59; Search similar-shape structures of this assembly by Omokage search (details)

Sample

• Sample: Name: Vaccinia virus A46 protein (full-length) / Specimen concentration: 4.4 mg/ml
• Buffer: Name: 20 mM Tris-HCl, 10 mM DTT / pH: 8.5

Entity #461

Name: VACV A46 / Type: protein / Description: Protein A46 / Formula weight: 28.019 / Num. of mol.: 4 / Source: Vaccinia virus / References: UniProt: P26672
Sequence:

GAQQMAFDIS VNASKTINAL VYFSTQQNKL VIRNEVNDTH YTVEFDRDKV VDTFISYNRH NDTIEIRGVL PEETNIGCAV NTPVSMTYLY NKYSFKLILA EYIRHRNTIS GNIYSALMTL DDLAIKQYGD IDLLFNEKLK VDSDSGLFDF VNFVKDMICC DSRIVVALSS LVSKHWELTN KKYRCMALAE HISDSIPISE LSRLRYNLCK YLRGHTESIE DKFDYFEDDD SSTCSAVTDR ETDV

Experimental information

• Beam: Instrument name: ESRF BM29 / City: Grenoble / 国: France / Type of source: X-ray synchrotronSynchrotron / Wavelength: 0.09918 Å / Dist. spec. to detc.: 2.867 mm
• Detector: Name: Pilatus 1M

Scan

Title: Vaccinia virus A46 protein (full-length) / Measurement date: Jun 25, 2015 / Cell temperature: 20 °C / Exposure time: 0.1 sec. / Unit: 1/nm /

	Min	Max
Q	0.1221	4.9336

Distance distribution function P(R)

Sofotware P(R): GNOM 5.0 / Number of points: 420 /

	Min	Max
Q	0.126754	2.09747
P(R) point	1	420
R	0	14

Result

Type of curve: single_conc /

	Experimental	Porod
MW	113.714 kDa	113.714 kDa
Volume	-	199 nm3

	P(R)	P(R) error	Guinier	Guinier error
Forward scattering, I0	75.37	0.116	75.53	0.11
Radius of gyration, Rg	4.32 nm	0.008	4.27 nm	0.08

	Min	Max
D	-	14
Guinier point	1	33