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Figure 4 | Virology Journal

Figure 4

From: Influenza A H3N2 subtype virus NS1 protein targets into the nucleus and binds primarily via its C-terminal NLS2/NoLS to nucleolin and fibrillarin

Figure 4

The C-terminal NoLS of the NS1 protein in the H3N2 subtype influenza A viruses binds in vitro -translated nucleolar nucleolin and fibrillarin in a GST pull-down experiment. [35S]-labeled and in vitro-translated nucleolin, B23, fibrillarin and, as a control, lamin A/C were allowed to bind to E. coli-expressed and Sepharose-immobilized GST, GST-NS1 A/Udorn/72 wt, GST-NS1 A/Udorn/72(1–73), GST-NS1 A/Udorn/72(1–73)R38A, GST-NS1 A/Udorn/72(1–73)K41A, GST-NS1 A/Udorn/72(203–237), GST-NS1 A/Udorn/72 (203–237)K219A,R220A + R231A,R232A, GST-NS1 A/Brevig mission/1/18 wt and GST-NS1 A/mallard/Netherlands/12/00 wt at +4 °C for 1 h. Sepharose-bound proteins were dissolved in Laemmli sample buffer, separated on 8 % SDS-PAGE and autoradiographed. Input (lane C) was 1:10 of the amount of [35S]-labeled protein that was used in the binding experiment. M indicates the Sepharose matrix-bound proteins. A similar gel was also stained with Coomassie Blue to visualize the amount of Sepharose-immobilized GST and GST-NS1 fusion proteins.

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