Topography of dolichyl phosphate synthesis in rat liver microsomes. Transbilayer arrangement of dolichol kinase and long-chain prenyltransferase.
The topography of the dolichyl phosphate biosynthetic enzymes within the plane of rat liver microsomes was investigated by the use of two impermeant inhibitors of enzyme activity: trypsin and mercury-dextran. Mercury-dextran was found to inactivate over 50% of the activities of the CTP-dependent dolichol kinase and the long-chain prenyltransferase. Trypsin caused over 90% inactivation of the long-chain prenyltransferase and 60% inactivation of the dolichol kinase. In addition, the CTP-dependent dolichol kinase was inhibited over 90% by CDP applied externally to sealed microsomes. Inactivation of the dolichyl phosphate biosynthetic enzymes by the impermeant probes occurred under conditions where the mannose-6-phosphatase activity was highly latent. It was concluded that the active sites of these two enzymes are located on the external surface of the microsomal membranes and that dolichyl phosphate biosynthesis occurs asymmetrically on the cytoplasmic surface of the endoplasmic reticulum.
D000818:Animals; D001665:Binding Sites; D002463:Cell Membrane Permeability; D003593:Cytoplasm / Q000201:enzymology; D003911:Dextrans / Q000494:pharmacology; D004122:Dimethylallyltranstransferase / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D007425:Intracellular Membranes / Q000201:enzymology*; D008628:Mercury / Q000494:pharmacology; D008862:Microsomes, Liver / Q000201:enzymology*; D017830:Octoxynol; D010770:Phosphotransferases / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D017853:Phosphotransferases (Alcohol Group Acceptor); D011092:Polyethylene Glycols / Q000494:pharmacology; D051381:Rats; D013439:Sulfhydryl Reagents / Q000494:pharmacology; D013499:Surface Properties; D014166:Transferases / Q000302:isolation & purification*; D014357:Trypsin / Q000494:pharmacology
D003911:Dextrans; D013439:Sulfhydryl Reagents; D011092:Polyethylene Glycols; D017830:Octoxynol; D014166:Transferases; D004122:Dimethylallyltranstransferase; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C020087:dolichol kinase; D014357:Trypsin; D008628:Mercury