Structural basis of carbohydrate transfer activity by human UDP-GalNAc: polypeptide alpha-N-acetylgalactosaminyltransferase (pp-GalNAc-T10).
Mucin-type O-glycans are important carbohydrate chains involved in differentiation and malignant transformation. Biosynthesis of the O-glycan is initiated by the transfer of N-acetylgalactosamine (GalNAc) which is catalyzed by UDP-GalNAc:polypeptide alpha-N-acetylgalactosaminyltransferases (pp-GalNAc-Ts). Here we present crystal structures of the pp-GalNAc-T10 isozyme, which has specificity for glycosylated peptides, in complex with the hydrolyzed donor substrate UDP-GalNAc and in complex with GalNAc-serine. A structural comparison with uncomplexed pp-GalNAc-T1 suggests that substantial conformational changes occur in two loops near the catalytic center upon donor substrate binding, and that a distinct interdomain arrangement between the catalytic and lectin domains forms a narrow cleft for acceptor substrates. The distance between the catalytic center and the carbohydrate-binding site on the lectin beta sub-domain influences the position of GalNAc glycosylation on GalNAc-glycosylated peptide substrates. A chimeric enzyme in which the two domains of pp-GalNAc-T10 are connected by a linker from pp-GalNAc-T1 acquires activity toward non-glycosylated acceptors, identifying a potential mechanism for generating the various acceptor specificities in different isozymes to produce a wide range of O-glycans.
Kubota
Tomomi
T
Glycogene Function Team of Research Center for Glycoscience (RCG), National Institute of Advanced Industrial Science and Technology (AIST), Central 2, 1-1-1 Umezono, Tsukuba, Ibaraki 305-8568, Japan.
Shiba
Tomoo
T
Sugioka
Shigemi
S
Furukawa
Sanae
S
Sawaki
Hiromichi
H
Kato
Ryuich
R
Wakatsuki
Soichi
S
Narimatsu
Hisashi
H
D000595:Amino Acid Sequence; D002241:Carbohydrates / Q000737:chemistry*; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D006031:Glycosylation; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics; D008345:Manganese / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009077:Mucins / Q000737:chemistry; D009154:Mutation; D017350:N-Acetylgalactosaminyltransferases / Q000737:chemistry* / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D013379:Substrate Specificity; D014530:Uridine Diphosphate / Q000737:chemistry; D000097763:Polypeptide N-acetylgalactosaminyltransferase
D002241:Carbohydrates; D007527:Isoenzymes; D008345:Manganese; D009077:Mucins; D017350:N-Acetylgalactosaminyltransferases; D011994:Recombinant Proteins; D014530:Uridine Diphosphate; D000097763:Polypeptide N-acetylgalactosaminyltransferase