Subcellular Location of NADP+-Isocitrate Dehydrogenase in Pisum sativum Leaves 1

AUTOR(ES)

Randall, Douglas D.

RESUMO

The subcellular location of NADP+-isocitrate dehydrogenase was investigated by preparing protoplasts from leaves of pea seedlings. Washed protoplasts were gently lysed and the whole lysate separated on sucrose gradients by a rate-zonal centrifugation. Organelles were located by marker enzymes and chlorophyll analysis. Most of the NADP+-isocitrate dehydrogenase was in the soluble fraction. About 10% of the NADP+-isocitrate dehydrogenase was present in the chloroplasts as a partially latent enzyme. Less than 1% of the activity was found associated with the peroxisome fraction. NADP+-isocitrate dehydrogenase was partially characterized from highly purified chloroplasts isolated from shoot homogenates. The enzyme exhibited apparent Km values of 11 micromolar (NADP+), 35 micromolar (isocitrate), 78 micromolar (Mn2+), 0.3 millimolar (Mg2+) and showed optimum activity at pH 8 to 8.5 with Mn2+ and 8.8 to 9.2 with Mg2+. The NADP+-isocitrate dehydrogenase activity previously claimed in the peroxisomes by other workers is probably due to isolation procedures and/or nonspecific association. The NADP+-isocitrate dehydrogenase activity in the chloroplasts might help supply α-ketoglutarate for glutamate synthase action.

Documentos Relacionados

• The NADP+-isocitrate dehydrogenase gene (icd) is nitrogen regulated in cyanobacteria.
• Partial Purification and Characterization of NADP+-Isocitrate Dehydrogenase from Immature Pod Walls of Chickpea (Cicer arietinum L.) 1
• Purification and Characterization of Cytosolic NADP Specific Isocitrate Dehydrogenase from Pisum sativum1
• Purification and Characterization of Chloroplastic NADP-Isocitrate Dehydrogenase from Mixotrophic Tobacco Cells (Comparison with the Cytosolic Isoenzyme).
• NADP-Isocitrate Dehydrogenase from Pseudomonas nautica: Kinetic Constant Determination and Carbon Limitation Effects on the Pool of Intracellular Substrates