Initiation of O-glycan synthesis in IgA1 hinge region is determined by a single enzyme, UDP-N-acetyl-α-D-galactosamine:: Polypeptide N-acetylgalactosaminyltransferase 2

The hinge region of human immunoglobulin A1 (*IgA1) possesses multiple O-glycans, of which synthesis is initiated by the addition of GaINAc to serine or threonine through the activity of UDP-N-acetyl-alpha-D-galactosamine:polypeptide N-acetylgalactosaminyltransferases (pp-GaINAc-Ts). We found that six pp-GaINAc-Ts, ppGaINAc-T1, -T2, -T3, -T4, -T6, and -T9, were expressed in B cells, IgA-bearing B cells, and NCI-H929 IgA myeloma cells. pp-GaINAc-T activities of these six enzymes for a synthetic IgA hinge peptide, which has nine possible O-glycosylation sites, were examined using a reversed phase-high performance liquid chromatography, a matrix-assisted laser desorption ionization time of flight mass spectrometry, and peptide sequencing analysis. pp-GaINAc-T2 showed the strongest activity transferring GaINAe to a maximum of eight positions. Other pp-GaINAc-Ts exhibited different substrate specificities from pp-GaINAc-T2; however, their activities were extremely weak. It was reported that the IgA1 hinge region possesses a maximum of five O-glycans, and their amino acid positions have been determined. We found that pp-GaINAc-T2 selectively transferred GaINAc residues to the same five positions. These results strongly suggested that pp-GaINAc-T2 is an essential enzyme for initiation of O-linked glycosylation of the IgA1 hinge region.