Galactosylation of endogenous proteins from human platelets.
Human platelets have been shown to contain the enzyme glycoprotein:galactosyltransferase that catalyzes the transfer of galactose to an endogenous protein acceptor present in the platelet. Galactosylation of added ovalbumin also occurs. The activity was extracted with 30 mM Tris buffer (pH 7.5). The endogenous activity was enriched 1.4-fold (compared with the crude homogenate) in the fraction, 105,000 g pellet, and the exogenous enzyme was retained in the respective supernatant. The two galactosyltransferase activities showed proportionality to time, protein, and substrate concentration, and were identical in pH dependence and Mn+2 requirement. The effect of Triton X-100 (range 0-1.5%) in the assay system appeared to be different for both activities: with the optimum concentration of detergent (0.15%) the endogenous activity increased by 50% whereas the exogenous activity was augmented 5-fold. From a number of sugar nucleotides tested as glycosyl donor into the endogenous proteins, the optimum substrate was UDP-Glc (100%), followed by UDP-Gal (80%), GDP-Man (24%), UDP-Glc-NAc (21%), UDP-Xyl (19%), and ADP-Glc (5%). An appropriate exogenous acceptor for UDP-Glc as donor was not found. The different solubilization of galactosyl- and glucosyltransferase activities by Triton X-100 suggests that they are distinct enzymes. In addition, the exogenous galactosyltransferase activity achieved after the treatment was much higher (940%) than the endogenous (26%). It is suggested that these differences on both galactosyltransferases could reflect changes in the accessibility of the exogenous substrate to the enzyme.
Rodríguez
P
P
Laboratorio de Trombosis Experimental Centro de Biofisica y Bioquímica, Instituto Venezolano de Investigaciones Científicas, Caracas.
Bello
O
O
Tablante
A
A
Apitz-Castro
R
R
D001792:Blood Platelets / Q000378:metabolism*; D001798:Blood Proteins / Q000378:metabolism*; D005700:Galactosyltransferases / Q000378:metabolism*; D005964:Glucosyltransferases / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D010047:Ovalbumin / Q000378:metabolism; D012995:Solubility; D014532:Uridine Diphosphate Glucose / Q000032:analysis
D001798:Blood Proteins; D010047:Ovalbumin; D005700:Galactosyltransferases; D005964:Glucosyltransferases; D014532:Uridine Diphosphate Glucose