Kinetic studies of alkyl-dihydroxyacetone-phosphate (alkyl-glycerone-phosphate) synthase in peroxisomes of rat liver.

The properties of peroxisomal enzyme alkylglycerone-phosphate synthase were studied in highly purified peroxisome fractions of rat liver. The requirements for optimal enzyme activity: pH and composition of the reaction mixture, incubation time, and enzyme concentration were investigated, and kinetic studies performed employing both different long-chain fatty alcohols and acyl dihydroxyacetone phosphates as substrates. Activities of the synthase considerably higher as reported before were found in the peroxisome preparation, with alkylglycerone (alkyldihydroxyacetone) phosphate as the sole product of the exchange reaction. The kinetic studies revealed divergent properties of peroxisomal synthase with respect to the substrates involved. Whereas the substrate concentration versus reaction velocity plot for the fatty alcohols reflects Michaelis-Menten kinetic behavior, it displays a maximum followed by inhibition with regard to the acylglycerone phosphate. The enzyme accepts different acylglycerone phosphates without much specificity but it is most active with 9-cis-octadecenol.

Distribution of alkylglycerone-phosphate synthase in subcellular fractions of rat liver.

Subcellular fractions of rat liver were isolated by density-gradient centrifugation on a linear Metrizamide gradient and were assayed for marker enzymes of peroxisomes, lysosomes, microsomes and mitochondria. Alkylglycerone-phosphate synthase catalysing the formation of the ether bond in glycerolipids was also determined along the gradient. The enzyme was found to be enriched in the peroxisomal and the microsomal fractions thus, displaying a bimodal distribution pattern. Two reaction-products each, alkylglycerone phosphate and alkylglycerone were obtained in the enzymic assays performed, the ratio of which was clearly dependent upon the fraction employed. Alkylglycerone phosphate was mainly synthesized by the 'peroxisomal synthase', whereas an inverse proportion was observed assaying the microsomal counterpart. Furthermore, comparing the mean specific activities of both the enzymes the microsomal one was shown to be roughly twice as active in metabolizing 1-O-palmitoylglycerone 3-phosphate, simultaneously displaying a somewhat different sensitivity to NaF. These findings provide a first line of evidence, that two separate synthases, one in microsomes and another one in peroxisomes might be engaged in the biosynthesis of 1-O-alkyl-glycerolipids in rat liver.