Purification, properties and partial amino acid sequence of the blood-group-A-gene-associated alpha-3-N-acetylgalactosaminyltransferase from human gut mucosal tissue.
An alpha-3-N-acetylgalactosaminyltransferase that transfers N-acetylgalactosamine from UDP-N-acetylgalactosamine to H-active structures to form A determinants was purified to homogeneity from human gut mucosal tissue of blood-group-A subjects. The mucosa was homogenized, then treated with Triton X-100, and the solubilized enzyme was purified by affinity chromatography on UDP-hexanolamine-agarose and octyl-Sepharose CL-4B. Enzyme activity was recovered in 44% yield with a specific activity of approx. 7 mumol/min per mg. The only effective acceptor substrates for the transferase were those containing a subterminal beta-galactosyl residue substituted at the O-2 position with L-fucose. The purified enzyme had a weak capacity to transfer D-galactose from UDP-D-galactose to similar acceptors to make blood-group-B determinants. H.p.l.c. and SDS/PAGE analysis indicated an Mr of 40,000 for the purified enzyme. For the first time a partial amino acid sequence Xaa-Ser-Leu-Pro-Arg-Met-Val-Tyr-Pro-Gln-Ile-Ser?-Val-Leu was obtained for the N-terminal region of the soluble alpha-3-N-acetylgalactosaminyltransferase.
Navaratnam
N
N
Division of Immunochemical Genetics, M.R.C. Clinical Research Centre, Harrow, Middx., U.K.
Findlay
J B
JB
Keen
J N
JN
Watkins
W M
WM
D000017:ABO Blood-Group System; D000595:Amino Acid Sequence; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D005690:Galactose / Q000378:metabolism; D005700:Galactosyltransferases / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007413:Intestinal Mucosa / Q000201:enzymology*; D008345:Manganese / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017350:N-Acetylgalactosaminyltransferases; D017830:Octoxynol; D011092:Polyethylene Glycols; D013379:Substrate Specificity; D014531:Uridine Diphosphate Galactose / Q000378:metabolism
D000017:ABO Blood-Group System; D014531:Uridine Diphosphate Galactose; D011092:Polyethylene Glycols; D008345:Manganese; D017830:Octoxynol; D005700:Galactosyltransferases; D017350:N-Acetylgalactosaminyltransferases; C003742:UDPgalactosamine-galactose acetylgalactosaminyltransferase; D005690:Galactose