Fig. 3

Expression and surface display of AIDA-I fusion proteins in E. coli, at 37 and 25 °C. Western blot of whole-cell lysates showing AIDA-I fusions proteins. Lane 1: Marker, Lane 2: NmCsgG, Lane 3: NmMtrC, Lane 4: NmBamE, Lane 5: Nm putative lipoprotein, Lane 6: EcBamE, Lane 7: EcLolB, Lane 8: EcLptE and Lane 9: EcPal, at 37 °C (a) and at 25 °C (b). FLAG-tag specific antibodies were used for detection. FACS analysis of E. coli BL21DE3 (pET15b) expressing AIDA-I fused to the N. meningitidis lipoproteins: CsgG, MtrC, BamE and putative lipoprotein at 37 °C (c) and 25 °C (d) and the E. coli lipoproteins: LolB, LptE, Pal and BamE at 37 °C (e) and 25 °C (f) were incubated with monoclonal anti-FLAG antibody. The grey areas represent the fluorescence signals obtained with the control (BL21DE3-pET15b ∅). The coloured lines represent the fused forms of the lipoproteins. Panels C and D (heterologous lipoproteins): Purple: CsgG, Dark Red: MtrC, Light Blue: nmBamE, Dark Green: putative lipoprotein; Panels E and F (homologous lipoproteins): Dark Blue: BamE, Light Green: LolB, Orange: LptE, Light Red: Pal