HPLC analysis of N-linked oligosaccharides from control and patient's fibroblasts reveal a delay of trimming from Man9GlcNAc2 to Man8GlcNAc2 in MAN1B1-deficient patients.

Protein N-linked oligosaccharides of two different control (A) and one MAN1B1-deficient individuals (P2, B) were separated by HPLC after the pulse period (left panels) and 2 hours of chase (right panels). Depicted for both controls and P2 are two representative chromatograms from two independent experiments, with one set of experiments presented in the upper panel, and a second one in the lower panel. Symbols: M8, M9 indicate oligosaccharide species with 8 or 9 mannose residues and possessing two GlcNAc resides at their reducing ends. G1M9 indicates oligosaccharide species with 9 mannose and 1 Glc residues and possessing two GlcNAc residues at their reducing ends. (C) Relative peak areas of M8, M9, and G1M9 were quantified and the results expressed as the percentage of processing efficiency to M8. In the calculation, the trimming efficiency from M9 to M8, which corresponds to the difference between the percentage of M8 at T0 h and T2 h, was considered as 100%. The depicted values are the mean ± SEM of independent experiments. (D) Comparison of the mean processing efficacy of the three control cell lines (C) to the mean processing efficacy of all four MAN1B1-deficient cell lines (P). Data were analysed by using Student's t test. *** indicates a p value<0.001.