Dihydroxyacetone Phosphate Reductase in Plants

Abstract

In plant metabolism glycerol phosphate must be produced for the synthesis of various phospholipids, sulfolipids, galactolipids, and triglycerides. The metabolic reaction in leaf tissue of higher plants for glycerol phosphate synthesis is catalyzed by a DHAP2 reductase which has been partially purified from spinach leaves and castor bean endosperm, and which catalyzes the reduction of DHAP at pH 7.0 using NADH as the reductant (6,17 No. 12290. 2Abbreviations: DHAP, dihydroxyacetone phosphate; ME, mercaptoethanol; PVPP, polyvinylpolypyrrolidone. metabolism must be highly controlled, as is expected of compounds and enzymes at metabolic branch points.A number of free living algae produce glycerol as a major product of photosynthesis rather than accumulate starch or sucrose. Zooxanthellae, symbiotic, unicellular algae in the polyps of reef building corals and other marine invertebrates, excrete to their host up to 40% of their photosynthate as glycerol (14, 15).The halotolerant alga, Dunaliella tertiolecta (5,20) and Chiamydomonas (1 1)

Two Isoforms of Dihydroxyacetone Phosphate Reductase from the Chloroplasts of Dunaliella tertiolecta

“…After reports that leaf or Dunaliella chloroplasts have a DHAP reductase glyceride isoform for glycerol-P formation for lipid production (Gee et al, 1988a(Gee et al, , 1988c, it was assumed that the glycerol-P pool in the Dunaliella chloroplasts would be used for either glycerol or for glyceride (lipid) synthesis. However, the presence in the Dunaliella chloroplasts of two distinctly different isoforms of DHAP reductase further suggests suborganelle enzymic localization of glycerol-P synthesis, one for glycerol production and one for the synthesis of lipids.…”

“…Leaves of higher plants contain both a chloroplastic and a cytoplasmic DHAP reductase form (Gee et al, 1988a(Gee et al, , 1988b. The presence of DHAP reductase has also been reported in Dunaliella tertiolecta (Haus and Wegman, 1984;Marengo et al, 1985; and Dunaliella parva (Gimmler and Lotter, 1982), and the enzyme was considered to be localized in the chloroplast (Brown et al, 1982;Gimmler and Lotter, 1982).…”

“…The activity as the glyceride form i n leaf homogenates could not be measured until after DEAE cellulose chromatography, due to the remova1 of low mo1 wt inhibitors, such as fatty acids and other membrane components (Gee et al, 1988a(Gee et al, , 1988b. However, the Dunaliella osmoregulator form was not inhibited by fatty acids and membrane components (next sections), so the nature of the inhibitors in the Dunaliella extract remains unknown.…”

“…DEAE cellulose was DE-52 from Whatman Ltd. Thioredoxin was from Escherichia coli (CalBiochem). The reduction of thioredoxin by DTT before use and its preparation for the DHAP reductase assay has been described (Gee et al, 1988a;modification of Wolosiuk et al, 1980). The total Chl content was determined spectrophotometrically at 645 and 663 nm on an aliquot of cells, chloroplasts, or homogenate that had been extracted with 80% acetone (Arnon, 1949).…”

“…The DHAP reductase activity was assayed at 25OC in the reductase direction at pH 6.9 expressed as pmol NADH oxidized h-' mg-' Chl (Gee et al 1988a). The glycerol-P dehydrogenase (oxidation direction) activity was assayed at pH 9.5 with 20 or 50 IJ~M DL-glycerol phosphate and 2 mM NAD or NADP.…”