Isolation and characterization of a rough microsomal fraction from rat kidney that is enriched in lysosomal enzymes.
1. A special population of rough microsomal material (microsomes) rich in lysosomal acid hydrolases was separated by isopycnic centrifugation as a discrete fraction (RM(2)) from the bulk of rough microsomal material in rat kidney because of its greater density. 2. The specific activities of five acid hydrolases in the RM(2) fraction were approximately one-half those of a purified lysosomal (L) fraction and 10- to 30-fold greater than those of an ordinary rough microsomal (RM(1)) fraction. 3. These special rough microsomes have a distinctive ultrastructure and electron-cytochemical properties. Their cisternal content resembles the matrix of lysosomes in that it is electron-dense, osmiophilic and plumbophilic and gives a positive reaction for acid phosphatase activity. 4. Polyacrylamide-gel electrophoresis of soluble proteins from the L fraction resolved nine anionic glycoproteins, most of which exhibit acid hydrolase activities (Goldstone & Koenig, 1970, 1973; Goldstone et al., 1971a). The most anionic glycoprotein is the acidic lipoglycoprotein of the lysosomal matrix (Goldstone et al., 1970). 5. Polyacrylamide-gel electrophoresis of soluble proteins from the RM(2) fraction resolved two cationic glycoproteins with acid hydrolase activities (Goldstone & Koenig, 1973) and an anionic glycoprotein with the same electrophoretic mobility as the lysosomal lipoglycoprotein, but without its lipid constituents or capacity to bind the basic fluorochrome Acridine Orange. These constituents are considered to be the precursors of the lysosomal glycoproteins.
D000135:Acid Phosphatase / Q000032:analysis; D000818:Animals; D002499:Centrifugation, Density Gradient; D004589:Electrophoresis, Disc; D005696:Galactosidases / Q000032:analysis; D005952:Glucose-6-Phosphatase / Q000032:analysis; D005966:Glucuronidase / Q000032:analysis; D005969:Glutamate Dehydrogenase / Q000032:analysis; D006596:Hexosaminidases / Q000032:analysis; D006651:Histocytochemistry; D007668:Kidney / Q000166:cytology / Q000201:enzymology*; D008247:Lysosomes / Q000032:analysis / Q000201:enzymology*; D008297:Male; D008854:Microscopy, Electron; D008861:Microsomes / Q000032:analysis / Q000201:enzymology*; D051381:Rats; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet; D013429:Sulfatases / Q000032:analysis; D014503:Urate Oxidase / Q000032:analysis
D005969:Glutamate Dehydrogenase; D014503:Urate Oxidase; D000135:Acid Phosphatase; D005952:Glucose-6-Phosphatase; D013429:Sulfatases; D005696:Galactosidases; D006596:Hexosaminidases; D005966:Glucuronidase