Biosynthesis of glycerolipids via acyldihydroxyacetone phosphate.

Acyldihydroxyacetone phosphate was first discovered as a lipid rapidly labelled by ’’Pi or by [p”P]ATP in guinea-pig liver mitochondria (Hajra & Agranoff, 1967,1968). This rapid labelling was due to the dephosphorylation and rephosphorylation of endogenous acyldihydroxyacetone phosphate in mitochondria (Hajra et al., 1968). Acyldihydroxyacetone phosphate was shown to be biosynthesized by the direct acylation of dihydroxyacetone phosphate with long-chain acyl-CoA species (Hajra, 1968~). This enzyme, acyl-CoA-dihydroxyacetone phosphate acyltransferase (EC 2.3.1.42), was found in both the microsomal and mitochondria1 fractions of different mammalian tissues (LaBelle & Hajra, 19726). In mitochondria, the enzyme is mostly located on the outer membrane (Jones & Hajra, 1976). We have purified the enzyme from guinea-pig liver mitochondria approx. 100-fold by a combination of sodium cholate solubilization, (NH,),SO, fractionation, gel filtration and DEAE-cellulose chromatography (Jones & Hajra, 1976). The purified enzyme was activated by the addition of phospholipids, and, like the membrane-bound enzyme, was found to be specific for only long-chain saturated acyl-CoA species as substrate. The partially purified enzyme did not have any acyl-CoA-glycerol 3-phosphate acyltransferase activity. Acyldihydroxyacetone phosphate was shown to be the precursor of glycerolipids containing an ether bond. After the discovery that long-chain alcohols and triose phosphates are the possible precursors of glycerol ether lipids (Friedberg & Greene, 1967, 1968; Ellingboe & Karnovsky, 1967; Snyder et al., 1969), it was found that dihydroxyacetone phosphate reacts with long-chain alcohols in the presence of ATP and CoA to form 1-0-alkyl dihydroxyacetone 3-phosphate (alkyldihydroxyacetone phosphate) (Hajra, 1969; Wykle & Snyder, 1969). A study of the detailed mechanism of the reaction showed that an intermediate was formed from dihydroxyacetone phosphate in the presence of ATP and CoA which was then converted into alkyldihydroxyacetone phosphate by reacting with long-chain alcohols (Hajra, 1970~). This intermediate was identified as acyldihydroxyacetone phosphate. Acyldihydroxyacetone phosphate reacts directly with long-chain alcohols to form alkyldihydroxyacetone phosphate (Hajra, 1970b; Wykle et al., 1972; Friedberg &Heifetz, 1975), andCoA is not necessary for the reaction. It was initially found that ATP stimulated the reaction, but recently we found that the stimulation was due to the binding of Mg2+ by ATP in the reaction mixture. Mgz+ inhibits the formation of alkyldihydroxyacetone phosphate (P. A. Davis & A. K. Hajra, unpublished work). Using specifically labelled dihydroxyacetone phosphate, Friedberg et al. (1972) and Friedberg & Heifetz (1973) have shown that there is a stereospecific exchange of hydrogen between water and C-1 of dihydroxyacetone 3-phosphate during the formation of the alkyl ether bond. Friedberg & Heifetz (1975) have also shown that ’H from 3Hz0 is incorporated into alkyldihydroxyacetone phosphate during its formation from acyldihydroxyacetone phosphate and long-chain alcohols. We confirmed these results and showed directly, by making specifically labelled 1-acyl Ror S[l-3H]dihydroxyacetone 3-phosphate, that one of the hydrogens at C-1 (pro-R) of acyldihydroxyacetone phosphate was exchanged with water during the formation of the