Microsequencing and cDNA cloning of the Calvin cycle/OPPP enzyme ribose-5-phosphate isomerase (EC 5.3.1.6) from spinach chloroplasts.
Ribose-5-phosphate isomerase (RPI) catalyses the interconversion of ribose-5-phosphate and ribulose-5-phosphate in the reductive and oxidative pentose phosphate pathways in plants. RPI from spinach chloroplasts was purified and microsequenced. Via PCR with degenerate primers designed against microsequenced peptides, a hybridisation probe was obtained and used to isolate several cDNA clones which encode RPI. The nuclear-encoded 239 amino acid mature RPI subunit has a predicted size of 25.3 kDa and is translated as a cytosolic precursor possessing a 50 amino acid transit peptide. The processing site of the transit peptide was identified from protein sequence data. Spinach leaves possess only one type of homodimeric RPI enzyme which is localized in chloroplasts and is encoded by a single nuclear gene. Molecular characterization of RPI supports the view that a single amphibolic RPI enzyme functions in the oxidative and reductive pentose phosphate pathways of spinach plastids.
Martin
W
W
Institut für Genetik, Technische Universität Braunschweig, Germany.
Henze
K
K
Kellerman
J
J
Flechner
A
A
Schnarrenberger
C
C
D019747:Aldose-Ketose Isomerases; D000595:Amino Acid Sequence; D001483:Base Sequence; D015139:Blotting, Southern; D002238:Carbohydrate Epimerases / Q000737:chemistry / Q000145:classification / Q000235:genetics*; D002736:Chloroplasts / Q000201:enzymology / Q000235:genetics*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D018628:Gene Dosage; D017343:Genes, Plant; D008969:Molecular Sequence Data; D010427:Pentose Phosphate Pathway; D010446:Peptide Fragments / Q000737:chemistry; D010940:Plant Proteins / Q000235:genetics*; D016133:Polymerase Chain Reaction; D017421:Sequence Analysis / Q000379:methods*; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D018724:Spinacia oleracea / Q000201:enzymology / Q000235:genetics*
D018076:DNA, Complementary; D010446:Peptide Fragments; D010940:Plant Proteins; D002238:Carbohydrate Epimerases; D019747:Aldose-Ketose Isomerases; C022548:ribosephosphate isomerase