Acinetobacter Phage AP205 VLP Design

The Acinetobacter phage, AP205, displaying two SpyTags per VLP subunit, was designed as previously described.³¹ (link) In brief, the gene encoding the AP205 coat protein (Gene ID: 956335) had the SpyTag peptide sequence (AHIVMVDAYKPTK) fused to both the N- and C-terminus. Each SpyTag was separated from the coat protein by a flexible linker (GSGTAGGGSGS, N-terminus and GTASGGSGGSG, C-terminus). The Spytagged VLP was expressed in Escherichia coli One Shot® BL21 Star™ (DE3) cells (Thermo Scientific) and purified by density gradient ultracentrifugation using an Optiprep™ step gradient (23, 29 and 35%) (Sigma).

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Palladini A., Thrane S., Janitzek C.M., Pihl J., Clemmensen S.B., de Jongh W.A., Clausen T.M., Nicoletti G., Landuzzi L., Penichet M.L., Balboni T., Ianzano M.L., Giusti V., Theander T.G., Nielsen M.A., Salanti A., Lollini P.L., Nanni P, & Sander A.F. (2018). Virus-like particle display of HER2 induces potent anti-cancer responses. Oncoimmunology, 7(3), e1408749.

Publication 2018

Escherichia coli One Shot® BL21 Star™ (DE3) cells Optiprep™ step gradient

Acinetobacter Cells Density gradient ultracentrifugation Escherichia coli Gene Phage Protein Spytag peptide Subunit

Corresponding Organization : University of Copenhagen

Other organizations : Istituto Ortopedico Rizzoli, University of California, Los Angeles

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Variable analysis

independent variables

Displaying two SpyTags per VLP subunit of the Acinetobacter phage AP205

dependent variables

Purification of the Spytagged VLP by density gradient ultracentrifugation

control variables

Expression of the Spytagged VLP in Escherichia coli One Shot® BL21 Star™ (DE3) cells
Use of flexible linkers (GSGTAGGGSGS, N-terminus and GTASGGSGGSG, C-terminus) to separate the SpyTag from the coat protein

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

Annotations

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