PHYTANIC ACID ACTIVATION IN RAT-LIVER PEROXISOMES IS CATALYZED BY LONG-CHAIN ACYL-COA SYNTHETASE

In Refsum disease, disorders of peroxisome biogenesis, and rhizomelic chondrodysplasia punctata, pathological accumulation of phytanic acid results from impaired alpha-oxidation of this branched chain fatty aci d. Previous studies from this laboratory indicated that activation of phytanic acid to its CoA derivative precedes its alpha-oxidation in pe roxisomes. It was reported that this reaction is catalyzed by a unique phytanoyl-CoA synthetase in human peroxisomes. We wanted to determine whether phytanic acid activation in rats required long-chain acyl-CoA synthetase (LCS), very long-chain acyl-CoA synthetase (VLCS), or a di fferent enyzme. To test directly whether LCS could activate phytanic a cid, rat liver cDNA encoding this enzyme was transcribed and translate d in vitro. The expressed enzyme had both LCS activity (assayed with p almitic acid, C16:0) and phytanoyl-CoA synthetase activity; VLCS activ ity (assayed with lignoceric acid, C24:0) was not detectable. The rati o of phytanoyl-CoA synthetase activity to palmitoyl-CoA synthetase act ivity for LCS synthetized in vitro (-20%) was higher than that observe d in peroxisomes isolated from rat liver (5-10%), suggesting that the expressed enzyme contained sufficient phytanoyl-CoA synthetase activit y to account for all activity observed in intact peroxisomes. Further experiments were carried out to verify that phytanic acid was activate d by LCS in rat liver peroxisomes. Attempts to separate LCS from phyta noyl-CoA synthetase by chromatography on several matrices were unsucce ssful. Preparative isoelectric focusing revealed that phytanoyl-CoA sy nthetase and LCS had indistinguishable isoelectric points. Phytanoyl-C oA synthetase activity was inhibited by unlabeled palmitic acid but no t by lignoceric acid. Heat treatment inactivated both phytanoyl-CoA an d palmitoyl-CoA synthetase activities at similar rates, 5,8,11,14-Eico satetraynoic acid inhibited activation of phytanic acid and palmitic a cid in a parallel dose-dependent manner, whereas activation of lignoce ric acid was not affected. These data support our conclusion that rat liver LCS, an enzyme known to be present in peroxisomal membranes, has phytanoyl-CoA synthetase activity. -Watkins, P. A., A. E. Howard, S. J. Gould, J. Avigan, and S. J. Mihalik. Phytanic acid activation in I at liver peroxisomes is catalyzed by long-chain acyl-CoA synthetase.