Biomedical subjects
B E Wright
Publications and source records attributed to B E Wright.
Metabolic compartments in Dictyostelium discoideum.
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Cystoid macular edema of nonretinal vascular origin.
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Kinetic models of metabolism in intact cells, tissues, and organisms.
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Kinetic models of glycogen metabolism in normal rat liver, Morris hepatoma 7787 and host liver.
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Glycogen metabolism in normal liver and Morris hepatoma 7787 in meal-fed rats.
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Mechanisms of development and aging.
The view is expressed that an inappropriate emphasis and research is currently directed towards simplistic approaches to the problem of aging; it is concluded that an understanding of the biochemical mechanisms underlying the aging process must be sought using a SYSTEMS analysis, at a level of complexity and integration much greater than that encompassed by any theory thus far considered.
The tricarboxylic acid cycle in Dictyostelium discoideum. Metabolite concentrations, oxygen uptake and 14c-labelled amino acid labelling patterns.
Some aspects of tricarboxylic acid-cycle activity during differentiation and aging in Dictyostelium discoideum were examined. The concentrations of glutamate, aspartate, alanine, citrate, 2-oxoglutarate, succinate, fumarate, malate, oxaloacetate, pyruvate and acetyl-CoA were determined at four stages over the course of differentiation. The rate of O2 utilization was also determined over differentiation. In addition, experiments are described in which the specific radioactivities of citrate, 2-oxoglutarate, succinate, fumarate and malate were determined during a 30 min labelling of cells from the preculmination stage of development with [14C]glutamate, [14C]aspartate or [14C]alanine. A similar experiment was also performed with cells from the aggregation stage of development using [14C]glutamate.
The tricarboxylic acid cycle in Dictyostelium discoideum. A model of the cycle at preculmination and aggregation.
A preliminary model of tricarboxylic acid-cycle activity in Dictyostelium discoideum is presented. Specific-radioactivity labelling patterns of intra- and extra-mitochondrial pools are simulated by this model and compared with the experimental data. The model arrived at by this method shows the following features. (1) The cycle flux rate is approx. 0.4 mM/min. (2) Both fumarate and malate are compartmentalized at approx. 1:5 between cycle pools and non-cycle pools. These may represent mitochondrial and cytoplasmic pools. Citrate is compartmentalized at 1:10. Succinate appears to exist in three compartments, two of which become labelled by [14C]glutamate and only one by [14C]aspartate (3) Two pools of aspartate with two associated pools of oxaloacetate are necessary for simulation. (4) Exchange between the cycle and non-cycle pools of both citrate and fumarate occurs at very low rates of about 0.003 mM/min, whereas exchange between the malate pools is about 0.004 mM/min. The exchange reaction glutamate in equilibrium 2-oxoglutarate runs at approx. 15 times the cycle flux. (5) A reaction catalysed by "malic" enzyme is included in the model, as this reaction is necessary for complete oxidation of amino acid substrates. (6) Calculation of the ATP yield from the model is consistent with earlier estimates of ATP turnover if the activity of adenylate kinase is considered.
The effects of exogenous glucose, uracil, and inorganic phosphate on differentiation in Dictyostelium discoideum.
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TFLUX: a general purpose program for the interpretation of radioactive tracer experiments.
TFLUX is a computer program which simulates radioactive tracer experiments using intact cells under steady-state conditions. It is designed for the naive computer user to use, providing English-like input. An example is described in which compartmentation and flux through the citric acid cycle in Dictyostelium discoideum are simulated.
Glycogen phosphorylase from Dictyostelium: a kinetic analysis by computer simulation.
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Amino acid catabolism and malic enzyme in differentiating Dictyostelium discoideum.
Amino acids produced from protein degradation are the major energy source for differentiation and aging in Dictyostelium discoideum. Considering the reactions involved in the conversion of amino acids from an average protein into tricarboxylic acid cycle intermediates, a route from a cycle intermediate (probably malate) to acetyl coenzyme A is required for the complete utilization of amino acids. Citrate was isolated from cells pulse-labeled with (14)C-labeled amino acids and was cleaved with citrate lyase. When cells were pulse-labeled with [U-(14)C]-glutamate the specific radioactivity of the acetate and oxaloacetate portions of citrate were consistent with the conclusion that one-third of the carbon flowing through the tricarboxylic acid cycle is removed for the synthesis of acetyl coenzyme A. The data were also consistent with the patterns of carbon flux required to maintain steady-state levels of cycle intermediates in cells catabolizing amino acids. It is suggested that the malic enzyme (EC 1.1.1.40) catalyzes the synthesis of acetyl coenzyme A from malate and is responsible for the observed citrate labeling pattern. In cell extracts the activity of this enzyme increased markedly with the onset of differentiation. The properties of partially purified (40-fold) malic enzyme isolated at culmination indicated that the enzyme was allosteric and was positively affected by aspartate and glutamate. Thus, amino acid production from protein degradation would stimulate a reaction essential for the efficient utilization of these amino acids for energy.
Effect of intracellular carbohydrates on heat resistance of Dictyostelium discoideum spores.
The effect of intracellular trehalose and glycogen on the survival of spores of Dictyostelium discoideum ATCC 25697 after exposure to supraoptimal temperatures was examined. Cells metabolically perturbed by incubation in glucose and inorganic phosphate have intracellular trehalose and glycogen concentrations fivefold and twofold higher, respectively, than those of the controls. These cells were more resistant to the lethal effects of wet heat (45 degrees to 55 degrees C) than were control cells. The presence of 40 mM trehalose in the buffer during heat stress increased the survival of nonperturbed cells to approximately the level of the perturbed cells. No protection was observed when cells were heated in the presence of exogenous glycogen. Glucose or disaccharides other than trehalose when present during heat stress, had no effect on heat resistance. Nonperturbed cells preincubated in 40 mM trehalose and washed before heat stress were more resistant to killing than were controls. Cells perturbed with inorganic phosphate, which has been shown to increase trehalose concentrations but decrease glycogen concentrations, were also more resistant to the lethal effects of wet heat than were controls. The data suggest that trehalose has an effect on the wet-heat resistance of D. discoideum. Some possible mechanisms are suggested.
A kinetic analysis of glucokinase and glucose-6-P phosphatase in Dictyostelium.
Using a mathematical model of carbohydrate metabolism in Dictyostelium discoideum, the kinetic expressions describing the activities of glucokinase and glucose-6-P phosphatase have been analyzed. The constraints on the kinetic mechanisms and relative activities of these two enzymes were investigated by comparing computer simulations to experimental data. The results indicated that, (1) glucose-6-P is compartmentalized with respect to the enzymes involved in glucose-60P, trehalose and glycogen metabolism, (2) a differences of approximately 0.6 mM/min in maximum specific activity of glucokinase compared to glucose-6-P phosphatase is required in order for the model to produce end product carbohydrate levels consistent with those observed experimentally, (3) the Km of glucokinase for glucose strongly influences the steady state levels of glucose in the absence of external glucose, and (4) changing the order of product removal in the reaction catalyzed by glucose-6-P phosphatase influences the level of glycogen and trehalose.
Kinetics of net RNA degradation during development in Dictyostelium discoideum.
Based on dry weight, first-order kinetics adequately describe the net decrease of RNA in Dictyostelium discoideum NC-4 until culmination, when a cessation of RNA degradation occurs. Between aggregation and sorocarp formation, the cells lose 40% of their RNA, or 19 mM-glucose equivalents expressed in terms of cell volume at aggregation. In sorocarps the spores contain a disproportionately large amount of RNA. Net RNA contents in sorocarps and in spores are unaffected by the availability of exogenous glucose. The relationship of dry weight, cell number and packed cell volume during differentiation is given.
Multiple forms of glucokinase from Dictyostelium discoideum.
A single form of glucokinase with an apparent Km value equal to 0.12 mM glucose was detectable in extracts prepared from aggregating cells, whereas kinetic and electrophoretic evidence indicated the presence of this form as well as a second glucose-phosphorylating enzyme with a Km value of about 0.01 mM glucose in extracts from culminating cells.