LC-MS/MS analysis of Mannose 6-phosphate containing proteins

Mucolipidosis III gamma (MLIII) is clinically characterized by onset of first symptoms at an average of 5 years such as stiffness of hands and shoulders, claw hand deformities, scoliosis and progressive destruction of hip joints. The disease is caused by mutations in GNPTG encoding the gamma-subunit of the GlcNAc-1-phosphotransferase complex. This enzyme is responsible for the generation of mannose 6-phosphate (M6P) targeting signals on 70 soluble lysosomal enzymes that are required for their efficient receptor-mediated transport to lysosomes. In fibroblasts from Gnptg-KO mice the GlcNAc-1-phosphotransferase activity is reduced leading to low amounts of M6P-containing proteins. M6P affinity chromatography-assisted and mass spectrometry-based secretome analysis of Gnptg-KO fibroblasts revealed a distinct set of lysosomal enzymes modified with M6P residues.