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 chromatography, 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 Thr4, Thr-3, then Thr-2 for UDP-N-acetyl-D-galactosamine:peptide N-acetylgalactosaminyltransferase-T3.
Iida
S
S
Laboratory of Cancer Biology and Molecular Immunology, Graduate School of Pharmaceutical Sciences, The University of Tokyo, Japan.
Takeuchi
H
H
Hassan
H
H
Clausen
H
H
Irimura
T
T
D000116:Acetylgalactosamine / Q000378:metabolism*; D000595:Amino Acid Sequence; D014408:Biomarkers, Tumor; D002458:Cell Fractionation; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D008861:Microsomes; D008969:Molecular Sequence Data; D055262:Mucin-2; D009077:Mucins / Q000138:chemical synthesis / Q000378:metabolism*; D017350:N-Acetylgalactosaminyltransferases / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000138:chemical synthesis / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid; D013379:Substrate Specificity; D013912:Threonine / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D000097763:Polypeptide N-acetylgalactosaminyltransferase
D000116:Acetylgalactosamine; D014408:Biomarkers, Tumor; D007527:Isoenzymes; D055262:Mucin-2; D009077:Mucins; D017350:N-Acetylgalactosaminyltransferases; D009363:Neoplasm Proteins; D011993:Recombinant Fusion Proteins; D013912:Threonine; D000097763:Polypeptide N-acetylgalactosaminyltransferase; C524625:MUC2 protein, human