FIG. 4.

Display of HIV Nef. Nef sequence was fused in frame to the N terminus of Hoc (A) and overexpressed in E. coli and purified (B). Lanes: 1 and 2, Nef-Hoc clones analyzed by SDS-10% PAGE before (0 h) or after (3 h) IPTG induction, respectively; 3, purified Nef-Hoc protein after Ni-affinity chromatography. (C) The hoc⁻soc⁻ particles (10¹⁰ PFU) were incubated with the purified Nef-Hoc at various ratios of Nef-Hoc to Hoc binding sites under standard in vitro assembly conditions. The samples were electrophoresed on an SDS-4 to 20% PAGE gel and stained with Coomassie blue. Lanes: 1, control hoc⁻soc⁻ phage; 2, 4, 6, 8, 10, and 12, Nef-Hoc at the ratios shown at the top; 3, 5, 7, 9, 11, and 13, phage showing displayed Nef-Hoc. (D) Table showing the binding parameters for Nef-Hoc calculated from C, as described in the legend to Fig. 3.