Figure 5.
Cellular localization of SpGH29C. A and B, activity of different cellular fractions of WT TIGR4 (A) and ΔspGH29C (B) against LewisX as detected by fluorophore-assisted carbohydrate electrophoresis. The activities of recombinant SpGH29C and BgaA against LewisX are shown as controls, and fucose is shown as a standard. C, background labeling of the different cellular fractions in the absence of LewisX. LewisX and LewisX treated with total soluble protein are shown for comparison. LeX, LewisX; EF, extracellular fraction; CF, cytoplasmic fraction; MF, membrane fraction. The 8-aminonaphthalene-1,3,6-trisulfonic acid (ANTS) lane indicates background labeling due to the fluorophore alone. Due to the background labeling of the cell wall–associated fraction, SpGH29C activity can be observed as a disappearance of LewisX rather than an appearance of fucose.