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Chloroplastic Glyceride Isoform of Dihydroxyacetone Phosphate Reductase from Dunaliella tertiolecta: Purification and Characterization

  • Durba Ghoshal
  • David Mach
  • Shilpa Gupta
  • Arun Goyal
Article

Abstract

The chloroplastic glyceride isoform of dihydroxyacetone phosphate reductase (Gly-DHAPR) in the photosynthetic unicellular green algae, Dunaliella, plays key role in the synthesis of glycerol-P and glycerides. A four-step procedure has been developed to purify the Gly-DHAPR from the chloroplasts of Dunaliella tertiolecta. The enzyme was purified 462-fold to apparent electrophoretic homogeneity by precipitation of Rubisco by polyethylene glycol-4000, and successive chromatography on DEAE cellulose, Sephacryl S-200, and Red Agarose. The overall yield of the purified enzyme was 5.1% with a specific activity of 425 μmol. min−1. mg−1 protein, and a subunit molecular mass of 37 kD. The Gly-DHAPR had little preference for NADH or NADPH, but was highly specific for DHAP. The purified enzyme was slightly stimulated by 50 mM NaCl, KCl or by 25 mM MgCl2. Detergents, lipids, fatty acids, or long-chain acyl-CoA derivatives inhibited the Gly-DHAPR. The Gly-DHAPR differs in properties from the other chloroplastic osmoregulatory isoform of DHAP reductase from Dunaliella, but has significant similarities with the glyceride isoforms from higher plants for glycerol-P and triglyceride synthesis.

Key words

dihydroxyacetone phosphate reductase Dunaliella tertiolecta Gly-DHAPR Glycerol-P Glycerol-P dehydrogenase Osm-DHAPR 

Abbreviations

Gly-DHAPR

Chloroplastic glyceride isoform of dihydroxyacetone phosphate reductase

Osm-DHAPR

Chloroplastic osmoregulatory isoform of dihydroxyacetone phosphate reductase

PEG

Polyethylene glycol-4000

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Copyright information

© Springer 2001

Authors and Affiliations

  • Durba Ghoshal
    • 1
  • David Mach
    • 1
  • Shilpa Gupta
    • 1
  • Arun Goyal
    • 1
  1. 1.Department of Biology, College of Science & Engineering, and Department of Biochemistry and Molecular Biology, School of MedicineUniversity of Minnesota DuluthDuluthUSA

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