Incorporation of N-acetylgalactosamine into consecutive threonine residues in MUC2 tandem repeat by recombinant human N-acetyl-D-galactosamine transferase-T1, T2 and T3

An oligopeptide containing three consecutive Thr residues mimicking the tandem repeat portion of MUC2. (PTTTPLK) was investigated for the acceptor specificity to UDP-N-acetyl-D-galactosamine:peptide N-acetylgalactosaminyltransferase isozymes, UDP-N-acetyl-D-galactosamine:peptide N-acetylgalactosaminyltransferase-T1, T2 and T3, The enzymatic reaction products were fractionated by the reversed-phase high performance liquid chromatograph?:, then characterized by matrix-assisted laser desorption ionization time of flight mass spectrometry and by a peptide sequencing analysis. A maximum of two, one or three N-acetyl-D-galactosamine residues was transferred by UDP-N-acetyl-D-galactosamine: peptide N-acetylgalactosaminyltransferase-T1, T2 or T3, respectively. The preferential orders of N-acetyl-D-galactosamine incorporation were Thr-2, then Thr-4 for UDP-N-acetyl-D-galactosamine :peptide N-acetylgalactosaminyltransferase-T1, Thr-2 for UDP-N-acetyl-D-galactosamine :peptide N-acetylgalactosaminyltransferase-T2 and Thr-4, Thr-3, then Thr-2 for UDP-N-acetyl-D-galactosamine :peptide N-acetylgalactosaminyltransferase-T3. (C) 1999 Federation of European Biochemical Societies.