THE ACCEPTOR SUBSTRATE-SPECIFICITY OF PORCINE SUBMAXILLARY UDP-GALNAC-POLYPEPTIDE N-ACETYLGALACTOSAMINYLTRANSFERASE IS DEPENDENT ON THE AMINO-ACID-SEQUENCES ADJACENT TO SERINE AND THREONINE RESIDUES

The acceptor substrate specificity of a pure polypeptide N-acetylgalac tosaminyltransferase has been examined with synthetic polypeptides wit h sequences identical, or similar to those found in porcine mucin or h uman erythropoietin. The sequences adjacent to either threonine or ser ine markedly influence the formation of GalNAc-O-Thr and GalNAc-O-Ser. Examination of the mucin-like peptide VLGXXAV, where X is Thr, Ser, o r Ala, shows only Thr-containing peptides to be acceptors. The best su bstrate is formed when XX is TT. Peptides with XX as either AT or TA a re less effective and those with XX as either ST or TS are much less e ffective acceptors. The amino acids adjacent to serine in the peptide formed by residues 121-131 in human erythropoietin, PPDAASAAPLR, also markedly influence the formation of GalNAc-O-Ser. Thus, PPDASSSAPLR an d PPDVVSVVPLR are about 5- and 30-fold, respectively, less active than the erythropoietin peptide. The peptide PPDGGSGGPLR is inactive. The shorter peptide DAASAAPL is also about 5-fold less active than the ful l-length peptide, but the peptide AASAA is inactive. These studies ind icate that one transferase can form both GalNAc-O-Ser and GalNAc-O-Thr residues when the sequences adjacent to the glycosylated residue are of the proper kind. Thus, in contrast to earlier suggestions, there is no evidence that different transferases form GalNAc-O-Ser and GalNAc- O-Thr. Examination of tissue homogenates from various tissues confirms this conclusion.