Alpha-hydroxylation of lignoceroyl-CoA by a cyanide-sensitive oxygenase in rat brain microsomes.
The enzymatic mechanism of alpha-hydroxylation of lignoceroyl-CoA, an intermediate in the synthesis of hydroxyceramide, was studied. In the presence of NADPH, sphingosine and microsomes from 20-day-old rat brain, 14C from [1-14C]lignoceroyl-CoA was incorporated into hydroxyceramide. Activity was linear with time (up to 40 min) and with protein (up to 0.8 mg). The apparent Km for lignoceroyl-CoA was about 10 microM. NADPH was a more efficient electron donor than NADH. Oxygen was required for activity, which increased linearly up to 20% O2. In 5 and 10% oxygen, the reaction was inhibited by 0.1 mM cyanide and by electron transfer chain inhibitors, cytochrome c, ferricyanide, menadione, and p-chloromercuriphenyl sulphonate; CO and SKF-525A had no effect. Moreover none of the inhibitors affected the formation of hydroxyceramide. Lignoceroyl-CoA alpha-hydroxylase appears to be an oxygenase requiring NADPH and oxygen, which involves cyanide-sensitive enzyme.
D000214:Acyl Coenzyme A / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000201:enzymology*; D002248:Carbon Monoxide / Q000494:pharmacology; D003486:Cyanides / Q000494:pharmacology*; D006900:Hydroxylation; D008861:Microsomes / Q000201:enzymology*; D009243:NAD / Q000494:pharmacology; D009249:NADP / Q000494:pharmacology; D010100:Oxygen / Q000494:pharmacology; D010105:Oxygenases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D013110:Sphingosine / Q000378:metabolism
D000214:Acyl Coenzyme A; D003486:Cyanides; D009243:NAD; C018582:lignoceroyl-coenzyme A; D009249:NADP; D002248:Carbon Monoxide; D010105:Oxygenases; D013110:Sphingosine; D010100:Oxygen