Human ovarian cancer, lymphoma spleen, and bovine milk GlcNAc:beta1,4Gal/GalNAc transferases: two molecular species in ovarian tumor and induction of GalNAcbeta1,4Glc synthesis by alpha-lactalbumin.
Affinity Gel-UDP was utilized to purify GlcNAc:beta1,4Gal/GalNAc transferases (Ts) from human lymphoma spleen, ovarian tumor, and ovarian cancer sera. Mn(2+) was found to be an absolute requirement for activity. Two molecular species containing both beta1,4Gal/GalNAc-T activities were discernible when the purified ovarian tumor microsomal enzyme was subjected to Sephacryl S-100 HR column chromatography as well as native polyacylamide gel-electrophoresis. Acceptor specificity studies of the affinity-purified lymphoma spleen and ovarian tumor microsomal enzymes and the conventionally purified, as well as the cloned, bovine milk GlcNAc:beta1,4Gal-Ts using a number of synthetic acceptors showed that the beta(1,6)-linked GlcNAc moiety to alpha-GalNAc was the most efficient acceptor. As compared to the purified milk enzyme, the recombinant form exhibited sixfold GlcNAc:beta1,4 GalNAc-T activity and up to eightfold GlcNAc6SO3beta-:beta1,4Gal-T activity. Further, the recombinant enzyme catalyzed the transfer of GalNAc to the terminal beta-linked GlcNAc6SO3 moiety. Alpha-lactalbumin (alpha-LA) inhibited up to 85%, the transfer of Gal to the GlcNAc moiety linked either to Man or GlcNAc. On the contrary, alpha-LA had no significant influence on the transfer of GalNAc to the above acceptors. alpha-LA had no appreciable effect on the recombinant enzyme, except for the transfer of Gal or GalNAc to Glc. Both alpha- and beta-glucosides, as well as alpha-N-acetylglucosaminide, did not serve as acceptors.
Chandrasekaran
E V
EV
Department of Molecular and Cellular Biophysics, Roswell Park Cancer Institute, Elm and Carton Streets, Buffalo, NY 14263, USA.
Chawda
R
R
Piskorz
C
C
Locke
R D
RD
Ta
A
A
Sharad
G
G
Odunsi
K
K
Lele
S
S
Matta
K L
KL
D000818:Animals; D002417:Cattle; D004187:Disaccharides / Q000378:metabolism; D005260:Female; D006020:Glycopeptides / Q000378:metabolism; D006689:Hodgkin Disease / Q000201:enzymology*; D006801:Humans; D007768:Lactalbumin / Q000378:metabolism / Q000494:pharmacology; D008892:Milk / Q000201:enzymology*; D017350:N-Acetylgalactosaminyltransferases / Q000187:drug effects / Q000378:metabolism*; D010051:Ovarian Neoplasms / Q000201:enzymology*; D013154:Spleen / Q000378:metabolism; D017855:Transferases (Other Substituted Phosphate Groups) / Q000378:metabolism
D004187:Disaccharides; D006020:Glycopeptides; D007768:Lactalbumin; C042746:N-acetylgalactosaminyl-(1-4)-glucose; D017350:N-Acetylgalactosaminyltransferases; D017855:Transferases (Other Substituted Phosphate Groups); C063044:UDP-N-acetylgalactosamine-N-acetylgalactosaminylphosphotransferase