Biosynthesis of Long Chain Alcohols in Developing Rat Brain.

Enzymatic formation of long chain alcohols, the first step of the pathway for the biosynthesis of ether lipids, was studied in rat brain. A particulate fraction from developing brain was found to catalyze an NADPH-dependent conversion of long chain acyl CoA to alcohol. A simple procedure was devised to chemically synthesize the substrate {('14)C}palmitoyl CoA at high specific activity. The reduction of this radioactive substrate by NADPH to {('14)C}hexadecanol, catalyzed by brain acyl CoA reductase, was stimulated by the addition of bovine serum albumin. The stimulation was due to the inhibition of an active acyl CoA hydrolase present in the particulate fraction used, rather than to a direct effect upon the reductase. An optimum ratio of albumin was found, approximately stoichiometric with amount of acyl CoA added. This finding is consistent with the postulate that monomeric, rather than micellar acyl CoA, is the substrate form utilized by the reductase. Acyl CoA reductase was most active with the palmitoyl-, stearoyl-, and oleoyl CoA's, thus accounting for the observed occurrence of only the corresponding alkyl and alkenyl moieties in the brain ether lipids. Acyl CoA reductase was concluded to have an essential thiol since it was inhibited by thiol agents. The presence of both substrates protected the enzyme from inactivation, suggesting that this thiol was located at the active site. The stoichiometry and stereospecificity of the reduction was determined by using stereospecifically labeled {4-('3)H}NADPH. Two moles of hydrogen, both from the B side of NADPH, were found to be utilized per mole of long chain alcohol formed. By the inclusion of aldehyde trapping agents and from examination of {4-('3)H}NADPH labeling patterns, the reduction of acyl CoA to alcohol was shown to be catalyzed by a single enzyme, rather than by two separate enzymes with a freely diffusible aldehyde intermediate. However, a small amount of aldehyde was produced by this enzyme, and this was postulated to be the result of the slow decomposition of an unstable enzyme-bound intermediate. Although most lipid biosynthetic enzymes are localized to the endoplasmic reticulum, it was found that acyl CoA reductase is probably localized to the microperoxisomes in brain. Procedures were developed for the isolation of this organelle from developing brain. It was inferred that the acyl CoA reductase was on the external surface of the microperoxisomal membrane because of its sensitivity to proteolysis. Acyl CoA reductase and catalase, the marker enzyme for microperoxisomes, were found to follow a similar developmental pattern in brain. Both were most active from birth to 10 days of age, decreasing thereafter to low levels in the adult. It was concluded that acyl CoA reductase controls the composition of the brain ether lipids, and perhaps also their rate of biosynthesis. The localization of this enzyme to microperoxisomes was postulated to implicate a role for this organelle in membrane biogenesis.Ph.D.BiochemistryUniversity of Michiganhttp://deepblue.lib.umich.edu/bitstream/2027.42/158218/1/8116200.pd