Abstract Polypeptide GalNAc-transferases (ppGalNAc-Ts) are a family of enzymes that catalyze the initiation of mucin-type O-glycosylation. All ppGalNAc-T family members contain a common (QXW)3 motif which is present in R-type lectin group. Acetylation site K521 is part of the QKW motif of ß-trefoil in the lectin domain of ppGalNAc-T2. We used a combination of acetylation and site-directed mutagenesis approaches to examine the functional role of K521 in ppGalNAc-T2. Binding assays of non-acetylated and acetylated forms of the mutant ppGalNAc-T2K521Q to various naked and aGalNAc-glycosylated mucin peptides indicated that degree of interaction of lectin domain with aGalNAc depends on the peptide sequence of mucin. Studies of inhibitory effect of various carbohydrates on interactions of ppGalNAc-T2 with MUC1aGalNAc indicate that point K521Q mutation enhance the carbohydrate specificity of lectin domain for aGalNAc. K521Q mutation resulted in enzyme activity lower than that of wild-type ppGalNAc-T2, similar to acetylation of ppGalNAc-T2. We conclude that an acetylation site in QKW motif of the lectin domain modulates carbohydrate recognition specificity and catalytic activity of ppGalNAc-T2 for partially pre-glycosylated acceptors and certain naked peptide. Post-translational modifications of ppGalNAc-Ts, such as acetylation, may play key roles in modulating functions of R-type lectin domains in cellular homeostasis.
Biological Chemistry, 2013, Vol 394, Issue 1, p. 69-77