Golgi-enriched membrane fractions from rat brain and liver contain long-chain polyisoprenyl pyrophosphate phosphatase activity.
The subcellular distribution of polyisoprenyl pyrophosphate phosphatase activity has been examined in rat brain by assaying the release of 32Pi from [beta-32P]dolichyl pyrophosphate (Dol-P-P) as described previously (Scher,M.G. and Waechter, C.J. (1984) J. Biol. Chem., 259, 14580-14585). The highest specific activities of Dol-P-P phosphatase in rat brain were found in the Golgi-enriched light microsomal, synaptic plasma membrane and heavy microsomal fractions. A comparative analysis of the distribution of galactosyltransferase and dolichol kinase reveals that Dol-P-P phosphatase activity co-fractionates with galactosyltransferase activity, and that the high level found in the Golgi-enriched fraction is not due to cross-contamination with heavy microsomes. When beta-labelled C95 Dol-P-P and the C95 allylic polyisoprenyl pyrophosphate (Poly-P-P) were compared as substrates for the Golgi-enriched light microsomal and heavy microsomal fractions, similar Km values were calculated for the two pyrophosphorylated substrates for each membrane fraction. Based on these kinetic analyses, the enzyme(s) catalysing this reaction do not distinguish between substrates containing saturated or allylic alpha-isoprene units. When Dol-P-P phosphatase activity was assessed in submicrosomal fractions obtained from rat liver by two separate procedures, the highest specific activity was also detected in the Golgi-enriched fraction. While the specific activities for Dol-P-P phosphatase and sialyltransferase were in the relative order of Golgi greater than smooth endoplasmic reticulum (ER) greater than rough ER, the relative order of dolichol kinase was rough ER greater than smooth ER greater than Golgi.(ABSTRACT TRUNCATED AT 250 WORDS)
Wolf
M J
MJ
Department of Biochemistry, University of Kentucky College of Medicine, A.B. Chandler Medical Center, Lexington 40536-0084.
Rush
J S
JS
Waechter
C J
CJ
D000818:Animals; D001921:Brain / Q000201:enzymology*; D004288:Dolichol Phosphates / Q000378:metabolism; D006056:Golgi Apparatus / Q000201:enzymology*; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D010744:Phosphoric Monoester Hydrolases / Q000032:analysis*; D011106:Polyisoprenyl Phosphates / Q000494:pharmacology; D011122:Polyphosphates / Q000378:metabolism; D011755:Pyrophosphatases; D051381:Rats; D012717:Sesquiterpenes; D013347:Subcellular Fractions / Q000201:enzymology; D013379:Substrate Specificity; D013835:Thiamine Pyrophosphate / Q000494:pharmacology; D014530:Uridine Diphosphate / Q000494:pharmacology
D004288:Dolichol Phosphates; D011106:Polyisoprenyl Phosphates; D011122:Polyphosphates; D012717:Sesquiterpenes; C026406:dolichol pyrophosphate; D014530:Uridine Diphosphate; C004808:farnesyl pyrophosphate; D010744:Phosphoric Monoester Hydrolases; D011755:Pyrophosphatases; C023082:dolichyldiphosphatase; D013835:Thiamine Pyrophosphate