title: Catalytic and glycan-binding abilities of ppGalNAc-T2 are regulated by acetylation.
abstract: Post-translational acetylation is an important molecular regulatory mechanism affecting the biological activity of proteins. Polypeptide GalNAc transferases (ppGalNAc-Ts) are a family of enzymes that catalyze initiation of mucin-type O-glycosylation. All ppGalNAc-Ts in mammals are type II transmembrane proteins having a Golgi lumenal region that contains a catalytic domain with glycosyltransferase activity, and a C-terminal R-type ("ricin-like") lectin domain. We investigated the effect of acetylation on catalytic activity of glycosyltransferase, and on fine carbohydrate-binding specificity of the R-type lectin domain of ppGalNAc-T2. Acetylation effect on ppGalNAc-T2 biological activity in vitro was studied using a purified human recombinant ppGalNAc-T2. Mass spectrometric analysis of acetylated ppGalNAc-T2 revealed seven acetylated amino acids (K103, S109, K111, K363, S373, K521, and S529); the first five are located in the catalytic domain. Specific glycosyltransferase activity of ppGalNAc-T2 was reduced 95% by acetylation. The last two amino acids, K521 and S529, are located in the lectin domain, and their acetylation results in alteration of the carbohydrate-binding ability of ppGalNAc-T2. Direct binding assays showed that acetylation of ppGalNAc-T2 enhances the recognition to αGalNAc residue of MUC1αGalNAc, while competitive assays showed that acetylation modifies the fine GalNAc-binding form of the lectin domain. Taken together, these findings clearly indicate that biological activity (catalytic capacity and glycan-binding ability) of ppGalNAc-T2 is regulated by acetylation.
authors.affiliation: Centro de Investigaciones en Química Biológica de Córdoba, UNC-CONICET, Departamento de Química Biológica, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Argentina.
authors.forename: Natacha
authors.initials: N
authors.lastname: Zlocowski
authors.forename: Victor G
authors.initials: VG
authors.lastname: Sendra
authors.forename: Virginia
authors.initials: V
authors.lastname: Lorenz
authors.forename: Marcos A
authors.initials: MA
authors.lastname: Villarreal
authors.forename: Alberto
authors.initials: A
authors.lastname: Jorge
authors.forename: Yolanda
authors.initials: Y
authors.lastname: Núñez
authors.forename: Eric P
authors.initials: EP
authors.lastname: Bennett
authors.forename: Henrik
authors.initials: H
authors.lastname: Clausen
authors.forename: Gustavo A
authors.initials: GA
authors.lastname: Nores
authors.forename: Fernando J
authors.initials: FJ
authors.lastname: Irazoqui
mesh_terms: D000107:Acetylation; D000595:Amino Acid Sequence; D002384:Catalysis; D006801:Humans; D008969:Molecular Sequence Data; D017350:N-Acetylgalactosaminyltransferases / Q000737:chemistry* / Q000235:genetics; D011134:Polysaccharides / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D000097763:Polypeptide N-acetylgalactosaminyltransferase
chemical_list: D017350:N-Acetylgalactosaminyltransferases; D011134:Polysaccharides; D000097763:Polypeptide N-acetylgalactosaminyltransferase