title: Subcellular localization of mannosyl transferase and glycoprotein biosynthesis in castor bean endosperm.
abstract: Differential and sucrose density gradient centrifugation have shown that the mannosyl transferase present in germinating castor bean endosperm cells which catalyses the synthesis of mannosyl-phosphoryl-polyisoprenol is exclusively located in the endoplasmic reticulum membrane. This intracellular location was confirmed using both ribosome-denuded microsomes isolated in the presence of EDTA and rough-surfaced microsomes isolated in the presence of excess Mg(2+) added to maintain ribosome-membrane attachment. Separation of organelles following the incubation of crude particulate fractions with GDP[(14)C]mannose demonstrated that most of the mannolipid thus formed remained associated with the microsomal fraction. When organelles were isolated from intact tissue which had previously been incubated with GDP[(14)C]mannose, [(14)C]glycoprotein was found to be associated with other cellular fractions in addition to the microsomes, in particular the glyoxysomes. The kinetics of radioactive labelling of these organelles suggest that [(14)C]glycoprotein appears initially in the microsomal fraction and subsequently accumulates in the glyoxysomes. Subfractionation of isolated, [(14)C]glycoprotein-labelled glyoxysomes established that over 80% of the total radioactivity was present in the membrane, while sodium dodecyl sulphate-polyacrylamide gel electrophoresis of solubilized glyoxysomal membranes showed that the [(14)C]sugar moiety was associated with several, but not all, constituent polypeptides.
authors: Postgraduate School of Biological Sciences, University of Bradford, BD7 IDP, Bradford, U.K..
authors: R B
authors: RB
authors: Mellor
authors: J M
authors: JM
authors: Lord