Biochemical studies of rat liver Golgi apparatus. I. Isolation and preliminary characterization.
A method is described for the isolation of morphologically well-preserved Golgi apparatus from rat liver. The method is essentially the same as that of Morré et al. (Morré, D.J., Hamilton, R.L., Mollenhauser, H.H., Mahley, R.W., Cunningham, W.P., Cheetham, R.D., & Lequire, V.S. (1970) J. Cell Biol. 44, 484-491) except that mild cell disruption is achieved by means of a stainless-steel sieve. The average recoveries of protein and galactosyltransferase in the isolated fraction are about 6 mg from 10 g of perfused liver and about 35% from the homogenate, respectively. The preparation is virtually free from succinate-cytochrome c reductase, glucose-6-phosphatase, acid phosphatase, and 5'-nucleotidase. The Golgi fraction as well as its vesicular fragments is homogeneous upon isopycnic centrifugation in both sucrose and dextran density gradients. Their buoyant densities in sucrose are significantly higher than those in dextran, indicating that both forms of the organelle are closed systems which are impermeable to macromolecules. The galactosyltransferase activity of a freshly prepared Golgi fraction, measured with ovalbumin as galactosyl acceptor, is activated 26-fold by the addition of Triton X-100, whereas those of homogenized, sonicated, and aged preparations are only activated 2- to 4-fold.
D000818:Animals; D002458:Cell Fractionation / Q000379:methods; D002500:Centrifugation, Isopycnic; D006056:Golgi Apparatus / Q000201:enzymology / Q000648:ultrastructure; D008099:Liver / Q000648:ultrastructure; D008247:Lysosomes / Q000201:enzymology; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology; D009237:N-Acetyllactosamine Synthase / Q000378:metabolism; D051381:Rats; D013347:Subcellular Fractions / Q000201:enzymology
D009237:N-Acetyllactosamine Synthase