Characterization of the reaction of GDP-mannose with dolichol phosphate in liver membranes.
The Mn-2+ dependent mannosyl transfer reaction between GDP-[14-C]mannose and dolichol phosphate, which is catalyzed by liver membranes, could not be followed accurately with the existing assay systems. Thus, GDP-[14-C]mannose is hydrolyzed rapidly by a pyrophosphatase present in microsomal and Golgi fractions from liver cells. The rate of the hydrolysis is rapid enough to limit the extent of incorporation of [14-c]mannose into endogenous acceptors. AMP was an effective inhibitor of the pyrophosphatase in Golgi membranes, and protected GDP-mannose from metabolism in alternative pathways. In the presence of AMP it was possible accurately to follow the time course of synthesis of dolichol phosphate [14-c]mannose over short time periods. Even though the time course of the reaction was measured over 2 s intervals, no linear portion could be detected in plots of product formed versus time. The kinetics of synthesis did, however, fit an equation for a first-order kinetic process. The basis for the first-order kinetics seems related to the very small amounts of dolichol phosphate in membranes. The values of the first-order rate constant is dependent on the concentrations of GDP-mannose and Mn-2+ added to the assays.
D000249:Adenosine Monophosphate / Q000494:pharmacology; D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D004224:Diterpenes / Q000378:metabolism*; D004286:Dolichols / Q000378:metabolism*; D006155:Guanosine Diphosphate Mannose / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008345:Manganese / Q000494:pharmacology; D008433:Mathematics; D009702:Nucleoside Diphosphate Sugars / Q000378:metabolism*; D009943:Organophosphorus Compounds / Q000378:metabolism; D051381:Rats; D013997:Time Factors
D004224:Diterpenes; D004286:Dolichols; D009702:Nucleoside Diphosphate Sugars; D009943:Organophosphorus Compounds; D006155:Guanosine Diphosphate Mannose; D000249:Adenosine Monophosphate; D008345:Manganese