Figure 4 | Scientific Reports

Figure 4

From: Site Specific Modification of Adeno-Associated Virus Enables Both Fluorescent Imaging of Viral Particles and Characterization of the Capsid Interactome

Figure 4

Biotinylation of modified AAV9 capsid enables the identification of interacting proteins. (A) Schematic showing two different ways to identify AAV9 interactors; In method 1, AAV9 particles are labelled in HEK cells 48 hrs after being generated before the complexes are isolated with streptavidin-agarose and identified through LC/MS; In method 2, purified high-titer AAV9 particles which have been biotinylated are mixed with mouse brain lysate before pulldowns (B) Instant Blue stained gel with fractions from pulldown with method 2 show enrichment of virus in AAV9-138 lane only; FW = first wash; LW = last wash. (C) Immunoblotting of streptavidin pulldowns with an anti-AAV antibody reveal bands corresponding to VP1 and VP3 in the AAV9-138 that are absent in controls (D) Gene ontology (GO) analysis taken from functional protein interaction networks (http://string-db.org/) reveals importance of RNA-binding and the actin cytoskeleton.

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