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. 2007 Apr 9;177(1):29–37. doi: 10.1083/jcb.200611079

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Copyright © 2007, The Rockefeller University Press

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Figure 3. — Reduced transfer of biosynthetic M₅GN₂-PP-Dol by the T. cruzi OST. (A) Biosynthetic M₅GN₂-PP-Dol (b) and M₅GN₂-PP-Dol isomers (c–i) produced by mannosidase digestion of M₉GN₂-PP-Dol (a). GlcNAc residues are indicated by squares, α-1,2–linked mannose residues are indicated by red circles, and α-1,3– and α-1,6–linked mannose residues are indicated by open circles. (B) Glycopeptide products obtained in an OST endpoint assay (>95% conversion of OS-PP-Dol to OS-NYT) were resolved by preparative HPLC to isolate the M₅GN₂-NYT glycopeptide (left). HPLC resolution of the α-1,2 mannosidase digestion products derived from M₅GN₂-NYT (right). The M₃GN₂-NYT (M3) peak is derived from isomer b, the M₄GN₂-NYT (M4) peak is derived from isomers c–h, and the M₅GN₂-NYT (M5) peak corresponds to isomer i. (C) HPLC profiles of α-1,2 mannosidase digestion products derived from M₅GN₂-NYT synthesized by the E. histolytica and T. cruzi OST. Redigestion of the M4 peak with a-1,2 mannosidase did not yield smaller products (not depicted); hence, the initial digestion had gone to completion. (D) The distribution of the three isomer classes (2, 1, or 0 α-1,2–linked mannose residues) was calculated for the total M₅GN₂-PP-Dol pool (OS) and for M₅GN₂-NYT synthesized by the S. cerevisiae (Sc), T. cruzi (Tc), E. histolytica (Eh), and T. vaginalis (Tv) OST. Values for the OS, Sc, and Tc are means of two independent experiments; error bars designate one of two independent data points. The OS values are derived from two replicates of B.