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Figure 2 | Retrovirology

Figure 2

From: APOBEC3G-UBA2 fusion as a potential strategy for stable expression of APOBEC3G and inhibition of HIV-1 replication

Figure 2

APOBEC3G fused with UBA2 domain is more resistant to Vif-mediated degradation than APOBEC3G. A-a. HEK293 cells, which is APOBEC3G-negative, was co-transfected with 1.5 μg of Vif-carrying plasmid (Vif-VR1012) DNA and 6 μg of plasmid DNA that expresses untagged APOBEC3G (E), APOBEC3G-UBA2 (U) fusion protein or APOBEC3G-UBA2* mutant fusion protein (M), respectively. Forty-five hours post-transfection (p.t.), cell lysates were subject to SDS polyacryladmide gel electrophoresis and analyzed by Western blot analysis using monoclonal anti-APOBEC3G and anti-Vif antibodies. Level of protein loading was measured by anti-β-actin antibody. A-b. The intensity of APOBEC3G protein was determined by densitometry. Value of the relative intensity of APOBEC3G was calculated in relative to the untagged APOBEC3G (E) and adjusted based on the relative intensity of β-actin in each lane to that of the control (C). B. UBA2 fusion to APOBEC3G does not affect its binding to Vif. Myc-tagged Vif was pulled-down in different APOBEC3G-producing HEK293 cells by immunoprecipitation using anti-Myc antibody. Binding of different forms of APOBEC3G to Vif was detected by using anti-APOBEC3G and anti-Vif antibodies, respectively. SUP, supernatants; IP, immunoprecipitation.

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