FIGURE 8.

GPI modification tolerates extensive sequence variation in the ω site/linker region. A, artificial GPI signals consisting of the flexible ω site/linker consensus sequence NSTGSGSSGS followed by Ala/Leu or Ala/Val/Phe hydrophobic stretches as shown were fused to Lep and tested in the in vitro translation system. B, GPI anchoring persists upon deletion of the ω environment. The C termini of CD24, COBL9, and EFNA1 were fused directly to the Lep protein replacing the ω site/linker by an Asn-Ser-Thr glycosylation acceptor site. C, addition of four lysines (4K) to the C terminus of Lep-GPI signal fusions efficiently blocks GPI anchoring, shown here for EFNA5, EFNA2, and CD24, and in Fig. 2B for PPB1 and FOL1. GPI signals are shown in italics and glycan acceptor sites are underlined. In panels B and C, GPI-anchored proteins that can be distinguished because of modified electrophoretic mobility are depicted with one or two arrowheads (single and double glycosylated, respectively). Other symbols are as in previous figures.