Fig. 2

Fig. 2 Activity-based identification of Q-glycosylating enzymes (A) Fractionation scheme of QTGAL (left) and QTMAN (right). (B) Fractionation of rat QTGAL activity by heparin chromatography (top), Mono S (middle), and SEC (bottom). Gray bars indicate protein amount (mg). Red plots show QTGAL activity measured by radioactivity of 14C-galactose incorporation (heparin and Mono S) or by LC/MS of galQ nucleoside (SEC). Molecular masses of size markers are indicated above the SEC profile. Proteins in the SEC fractions are resolved by SDS-PAGE. The band indicated by a red arrowhead was assigned to QTGAL (B3GNTL1). (C) CID spectrum of a tryptic fragment of the excised protein band. b and y series product ions are assigned to the tryptic peptide of rat B3GNTL1. (D) Fractionation of pig QTMAN activity by heparin and Resource Q chromatographies. Gray bars indicate protein amount (mg). Green plots show QTMAN activity measured by LC/MS of manQ nucleoside. Proteins fractionated by Resource Q are resolved by SDS-PAGE. See also Figures S10 and S11.