Light-induced accumulation of lactate and succinate in Anabaena cylindrica

In the cyanobacterium Anabaena cylindrica lactate accumulated in large amounts when the cells were exposed to light. The presence or absence of oxygen, or a change in CO₂ concentration did not affect the lactate accumulation. The cellular succinate level also increased in the light when CO₂ was supplied at the high concentration of 1%. 3-(3,4-dichlorophenyl)-1,1-dimethyl urea (DCMU), an inhibitor of photosynthetic electron flow, inhibited the increase in the concentration of lactate and succinate. Photosynthesis is a prerequisite for the increase of these organic acids. Thenoyltrifluoroacetone, an inhibitor of succinate dehydrogenase, inhibited the increase of succinate, suggesting that the succinate is formed via fumarate by the reverse of reactions of tricarboxylic acid (TCA) cycle. Upon addition of ammonium to the cell suspension in the light under high CO₂ concentration, the increases in the concentrations of lactate and succinate were inhibited while those of glutamine, glutamate and aspartate were stimulated. Ammonium apparently changed the products of metabolism of pyruvate and oxaloacetate from lactate and succinate to amino acids.Abbreviations Chl chlorophyll - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl urea - TTFA thenoyltrifluoroacetone - PCA perchloric acid     

Studies on NADP-isocitrate dehydrogenase from maturing castor bean seeds

NADP-specific isocitrate dehydrogenase from the soluble fractionof maturing castor bean endosperm was partially purified (approximately180-fold) and some of its enzymatic properties were studied.Mg⁺⁺, Mn⁺⁺, Cd⁺⁺, Ba⁺⁺, Co⁺⁺, Zn⁺⁺, and Sr⁺⁺ were activatorsof the enzyme reaction at a concentration of 6.7x10 M. The optimumpH of this enzyme was about 8.5. The enzyme was stable in thenarrow range from pH 7.0 to pH 8.0. Km values for isocitrateand NADP at pH 8.5 were 3.5x10^–6 M and 3.6x10^–6M, respectively. Enzyme stability was not affected by NaCl concentrationand enzyme reaction was inhibited at 5x10^–6 M PCMB (80%inhibition). It is suggested that the condensation product ofglyoxylate and oxalacetate also inhibits the reaction. NADP-IDHin the crude extract from maturing castor bean endosperm washeat-stable but the dialyzed enzyme preparation and the partiallypurified enzyme were labile against heat treatment at 57°C.When Mg⁺⁺ was added to the partially purified enzyme in thepresence of isocitrate or NADP, the enzyme was stabilized againstheat treatment. Mn⁺⁺, Ca⁺⁺, Co⁺⁺, Sr⁺⁺ or Ba⁺⁺ could be substitutedfor Mg⁺⁺. Addition of only one of the factors, Mg⁺⁺, isocitrateor NADP, had no effect on the heat stability. Moreover, a combinationof isocitrate and NADP did not establish stabilization. A divalentcation plays a central role, while adenine nucleotide, especiallyATP, may have an important part in stabilization. (Received August 14, 1972; )     

Characteristics of the reverse reaction of NADP+-isocitrate dehydrogenase from castor bean seeds

The reductive carboxylation of α-ketoglutarate by purified NADP⁺-isocitrate dehydrogenase (EC 1.1.1.42) from maturing castor bean seeds ( Ricinus communis L. ) has been characterized. The optimum pH for the reaction was 6.5, whereas pH 8.5 was optimum for oxidation of isocitrate (forward reaction). The enzyme utilized NADH as well as NADPH as the reducing agent in the reverse reaction, but only NADP⁺ in the forward reaction. The Km values for NADPH and NADH were 0.044 and 2.8 m M respectively, and for α-ketoglutarate and HCO₃ 4.1 and 3.7 m M. The enzyme was activated by various cations including Mg²⁺, Mn²⁺, Co²⁺, Zn²⁺, Ni²⁺ and Co²⁺. Km values for Mg²⁺ Mn²⁺, Co²⁺ and Zn²⁺ were 12, 34, 37 and 49μ M respectively.     

Glucose-6-Phosphate Dehydrogenase in Plastids from Developing Castor Bean Seeds

Glucose-6-phosphate dehydrogenase, together with the other enzymesof pentose phosphate pathway, was found in the cytosol as wellas in the plastid from developing castor bean (Ricinus communisL.) seeds. The plastid enzyme was found in both the matrix andthe membrane. The plastid enzyme has a sharp pH profile withthe optimum at 8.5, while the cytosolic enzyme has a broad pHprofile, optimum at 7.5. The plastid enzyme was inactivatedby storage at 0°C and by detergents such as Triton X-100,Brij and Nonidet, but the cytosolic enzyme was not. Slab geldisc electrophoresis indicated that three isoenzymes of glucose-6-phosphatedehydrogenase were found in the plastid but one enzyme in thecytosol of developing castor bean seed. From the presence ofglucose-6-phosphate dehydrogenase in the plastid, the operationof whole pentose phosphate pathway in this organelle of developingcastor bean seeds is suggested. (Received September 21, 1982; Accepted January 17, 1983)