Fig. 4.
Elongation of preexisting pools of mannan. (A) L. mexicana Δpmi promastigotes were cultivated in RPMI medium 1640 supplemented with 200 μM Man and pulse–chase labeled with [3H]Man. Promastigotes were sampled at the indicated times during the chase in 200 μM Man, and polar metabolites were desalted before or after mild acid hydrolysis (+H). The labeled products were analyzed by HPTLC and detected by fluorography. (B) The neutral [3H]mannan fraction from A was hydrolyzed in TFA (0.2 M at 100°C) for 0, 1, and 2 h and then analyzed by HPTLC. Lanes 1–3 were stained with orcinol/H2SO4; lanes 4–6 were subjected to fluorography. (C) Incubation of particulate (p) and cytosolic (c) fractions from L. mexicana Δgmp promastigotes with buffer control (−) or unlabeled neutral β-1,2-mannan [(M)n] and GDP-[3H]Man.