Galactosyltransferase (GalTase) prepared from human milk was found to exist as a complex with e-lactalbumin as demonstrated by crossed immunoelectrophoresis against specific antibodies raised against the complex. GalTase activity was stable to proteolysis and, when subjected to gel filtration on Ultrogel AcA54, the enzyme activity eluted as a single peak. A second peak of activity was found to be adsorbed to the column matrix and was eluted with buffer containing 1 M NaC1. The hydrophobic fraction represented 5% of the total GalTase activity in human milk. After polyacrylamide gel electrophoresis the main enzyme activity peak was represented by polypeptides of 67kDa molecular weight and of 14kDa molecular weight. Electroblotting of these peptides onto a nitrocellulose membrane followed by determination of GalTase activity showed activity for 45–55 kDa and for 14 kDa peptides. The hydrophobic fraction from the AcA54 column was resolved into polypeptides of 110 kDa-45 kDa molecular weight, all of which contained GalTase activity after blotting. It is supposed that the GalTase from non-proteolyzed milk is composed of a 14 kDa polypeptide containing the active site together with another part of the polypeptide backbone which is involved in the regulation of GalTase activity by α-lactalbumin, a third part of the polypeptide is responsible for the membrane insertion.
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September 01 1987
A solid phase assay for galactosyl transferases. Evidence for an active site of less than 14 kDa
Yves Plancke;
Yves Plancke
1
1Université des Sciences et Techniques de Lille Flandres-Artois, Laboratoire de Chimie Biologique et Laboratoire Associé au CNRS No. 217, 59655 Villeneuve d'Ascq Cédex, France
1To whom correspondence should be addressed.
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Béatrice Delpouve;
Béatrice Delpouve
1Université des Sciences et Techniques de Lille Flandres-Artois, Laboratoire de Chimie Biologique et Laboratoire Associé au CNRS No. 217, 59655 Villeneuve d'Ascq Cédex, France
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Jean Montreuil
Jean Montreuil
1Université des Sciences et Techniques de Lille Flandres-Artois, Laboratoire de Chimie Biologique et Laboratoire Associé au CNRS No. 217, 59655 Villeneuve d'Ascq Cédex, France
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Publisher: Portland Press Ltd
Received:
August 20 1987
Online ISSN: 1573-4935
Print ISSN: 0144-8463
© 1987 Plenum Publishing Corporation
1987
Biosci Rep (1987) 7 (9): 721–729.
Article history
Received:
August 20 1987
Citation
Yves Plancke, Béatrice Delpouve, Jean Montreuil; A solid phase assay for galactosyl transferases. Evidence for an active site of less than 14 kDa. Biosci Rep 1 September 1987; 7 (9): 721–729. doi: https://doi.org/10.1007/BF01116865
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