Edith M. Lees

UREIDOGLYCOLATE AMIDOHYDROLASE FROM DEVELOPING FRENCH BEAN FRUITS (PHASEOLUS VULGARIS L..)

Ureidoglycolate is an intermediate of allantoin catabolism in ureide-transporting legumes. This report describes the first purification of ureidoglycolate degrading activity (UGDA) from plant tissue in which the enzyme has been separated from urease. The enzyme from developing fruits of Phaseolus vulgaris has been purified 48-fold to give a preparation free of allantoinase and urease activity. UGDA was inhibited by EDTA while the Vmax was increased in the presence of Mn2+. The Km values for ureidoglycolate in the presence and the absence of Mn2+ were 2.0 and 5.4 mM, respectively. In the absence of Mn2+ UGDA was heat labile at 40 degrees C, but in the presence of Mn2+ the activity was stable up to temperatures of 60 degrees C. The Mr of UGDA was determined to be 300,000 by gel filtration chromatography and the pH optimum ranged from pH 7.0 to 8.5. Ammonia was determined to be the nitrogen-containing product of UGDA by a microdiffusion assay. This enzyme should therefore be described as ureidoglycolate amidohydrolase. The activity was shown to be associated with peroxisomes by fractionation of a crude extract on a sucrose density gradient. The products of ureidoglycolate degradation are glyoxylate, ammonia, and presumably carbon dioxide, which can be readily utilized by pathways of metabolism that are known to be present in this organelle.

The distribution of asparaginase activity in legumes

Abstract Legumes have been examined for asparaginase activity, taking into consideration their symbiotic association with bacteria in their nitrogen-fixing nodules. It has been shown that legumes grown without these bacterial asociations can hydrolyse asparagine and that asparaginase is located principally in the roots of the plants. However, in a normally nodulated plant the non-bacteroid portion of the nodule has a high concentration of asparagine, and asparaginase activity is much higher per g of tissue in this part of the nodule than in the remainder of the root section of the plant. Bacteroids from the nodule are also able to hydrolyse asparagine. The possible contribution of the nodule to the asparagine metabolism of the whole plant is discussed.

Cadmium bioaccumulation in Proisotoma minuta in relation to bioavailability in soils

Proisotoma minuta are widely distributed in Australian soils, especially in rehabilitated mine sites and in cotton fields. Laboratory studies were conducted to evaluate the accumulation of cadmium (Cd) in the Collembolan, P. minuta, using four Australian soils. Cadmium accumulation in body tissues was measured by atomic absorption spectroscopy of acid digests of the Collembola. Cadmium soil bioavailability of four Australian soils was determined using a sequential extraction method. The highest soluble-exchangeable Cd concentration was observed in Box Hill soil with the lowest pH, organic carbon, CEC, clay and very low iron content. Robertson soil with high pH, CEC, organic carbon, clay and iron content had the highest Cd organic- and oxide-bound concentrations. This may explain why there was higher Cd accumulation in P. minuta in the Box Hill soil than in the Robertson soil. The Cd uptake of P. minuta was predominantly correlated with soluble-exchangeable Cd concentration in soils followed by organic- and oxide-bound Cd concentrations. Presumably, soluble-exchangeable Cd is more readily available for P. minuta uptake than other Cd fractions.

Glutamate dehydrogenase activity in developing soybean seed: Isolation and characterization of three forms of the enzyme☆

Abstract Three forms of glutamate dehydrogenase have been isolated from developing soybean seed. Their intracellular locations could not be determined directly because organelles and marker enzymes showed abnormal distribution on sucrose density gradient fractionation. By analogy with enzymes from other parts of the plant, glutamate dehydrogenase 2 was shown to be located in chloroplasts and glutamate dehydrogenase 3 in mitochondria. Glutamate dehydrogenase 1 could not be located in this way because it is found only in the seed. The three enzymes are similar in pH optima, molecular weight and substrate specificity with respect to 2-oxoglutarate and l -glutamate. The mitochondrial enzyme is specific for NAD+. The chloroplast enzyme shows low activity with NADP+ relative to NAD+ but uses NADPH readily in the aminating direction. Glutamate dehydrogenase 1 is active with both nucleotides and is the only form to show substantial deaminating activity with NADP+. Glutamate dehydrogenases 1 and 2 are activated and stabilized by glutathione and 2-mercaptoethanol whereas enzyme 3 is unaffected. No significant metabolic control of any of the enzymes could be detected. Malate, citrate, adenine nucleotides and long-chain fatty acyl CoA derivatives gave slight inhibition at high concentrations. Amino acids had no effect on activity. A possible role for the enzyme characteristic of the developing seed is discussed in relation to nutrient supply during the accumulation of reserve materials in the seed.

Glutamate dehydrogenase activity in developing soybean seed: Kinetic properties of three forms of the enzyme

Abstract The kinetic parameters of three forms of glutamate dehydrogenase from developing soybean seed have been determined. For all enzymes double reciprocal plots were intersecting in animating and deaminating directions when each substrate was held at a fixed concentration in turn. In amination all plots were linear and the KNH4+ and KNAD(P)H values determined using (NH4)2SO4 were about one-fifth of values obtained with NH4CL. In deamination, plots were linear when l -glutamate and NADP+ were varied and biphasic with varied NAD+. It was not possible to determine the mechanisms of the enzymes but the data are consistent with nucleotide binding to the free enzyme forms. The role of the enzymes in the tissue is discussed. They could be active in deamination in the later stages of seed development when the supply of carbohydrate could be limiting. They could also be important in ammonia assimilation and their kinetic properties are discussed in relation to known properties of glutamine synthetase and glutamate synthase and cellular levels of enzyme substrates.

Determination of Oxidative Metabolism in Collembolan Proisotoma minuta (Tullberg)

An oxidative metabolism of Collembolan Proisotoma minuta was determined with a model compound of aldrin and dieldrin in this paper. The seven-day LD50 values for aldrin, dieldrin, and piperonyl butoxide in salt solution were 0.496, 0.367, and 8.346 mg L−1, respectively. When P. minuta were exposed to aldrin, dieldrin was the sole metabolite. The conversion of aldrin to dieldrin was known to be catalyzed by P450 monooxygenases system. It has been shown that the synergist piperonyl butoxide inhibited the metabolism of aldrin in P. minuta.