FIG. 4.

Both glycolipids GL-4 and GL-5 serve as substrates for GlfT2. Natural substrates GL-4 and GL-5 were used as acceptors of galactofuranosyl residues in the reaction catalyzed by GlfT2 expressed in the heterologous host, E. coli. The reaction mixtures were subjected to series of extractions, as described. (A) TLC of glycolipid fractions followed by autoradiography. (B) SDS-PAGE of lipid-linked galactan polymer followed by Western blotting and autoradiography. Lanes labeled “control” represent samples containing the host E. coli strain. Lanes labeled “standard” represent fractions extracted from reaction mixture containing membranes and cell wall from M. smegmatis mc²155 and UDP-[U-¹⁴C]Galp as a tracer for production of galactan intermediates (25).