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Temperature and the regulation of enzyme activity in poikilotherms. Properties of rainbow-trout fructose diphosphatase.

ABSTRACT: 1. The properties of fructose diphosphatase from the liver of rainbow trout (Salmo gairdnerii) were examined over the physiological temperature range of the organism. 2. Saturation curves for substrate (fructose 1,6-diphosphate) and a cofactor (Mg(2+)) are sigmoidal, and Hill plots of the results suggest a minimum of two interacting fructose 1,6-diphosphate sites and two interacting Mg(2+) sites per molecule of enzyme. 3. Mn(2+)-saturation curves are hyperbolic, and the K(a) for Mn(2+), which inhibits the enzyme at high concentrations, is 50-100-fold lower than the K(a) for Mg(2+). 4. Fructose diphosphatase is inhibited by low concentrations of AMP; this inhibition appears to be decreased and reversed by increasing the concentrations of Mg(2+) and Mn(2+). Higher concentrations of AMP are required to inhibit the trout fructose diphosphatase in the presence of Mn(2+). 5. The affinities of fructose diphosphatase for fructose diphosphate and Mn(2+) appear to be temperature-independent, whereas the affinities for Mg(2+) and AMP are highly temperature-dependent. 6. The pH optimum of the enzyme depends on the concentrations of Mg(2+) and Mn(2+). In addition, pH determines the K(a) for Mg(2+); at high pH, K(a) for Mg(2+) is lowered. 7. The enzyme is inhibited by Ca(2+) and Zn(2+), and the inhibition is competitive with respect to both cations. 8. The possible roles of these ions and AMP in the modulation of fructose diphosphatase and gluconeogenic activity are discussed in relation to temperature adaptation.

SUBMITTER: Behrisch HW

PROVIDER: S-EPMC1187510 | biostudies-other | 1969 Feb

REPOSITORIES: biostudies-other

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Project description:1. The properties of fructose diphosphatase from skeletal muscle of the Alaskan king-crab (Paralithodes camtschatica) were examined over the physiological temperature range of the animal. 2. King-crab muscle fructose diphosphatase is first activated by Na(+) and NH(4) (+) and is then partially inhibited by these cations at concentrations higher than 10mm at 0 degrees , 8 degrees and 15 degrees C. Enzyme activity is stimulated by K(+) at 0 degrees C, but is curtailed at 8 degrees C and 15 degrees C, an effect that could render rate independent of temperature. 3. Affinity for substrate increases with decreasing temperature; below the temperature of acclimatization, K(m) for fructose 1,6-diphosphate increases, resulting in a complex U-shaped temperature-K(m) curve. 4. King-crab muscle fructose diphosphatase is inhibited by low concentrations of AMP. As with enzymes of other poikilotherms, inhibition by AMP is sensitive to temperature; the enzyme is least sensitive to inhibition by AMP near the temperature of acclimatization. 5. The affinity of fructose diphosphatase for fructose 1,6-diphosphate is enhanced by phosphoenolpyruvate, and this activation is temperature-sensitive; 0.5mm-phosphoenolpyruvate causes a sevenfold decrease in K(m) for fructose 1,6-diphosphate at 15 degrees C but a 25-fold decrease at 0 degrees C. 6. Phosphoenolpyruvate appears to decrease the affinity of king-crab muscle fructose diphosphatase for AMP at low temperature, whereas at the higher temperature it appears to enhance inhibition by AMP. Phosphoenolpyruvate was not observed to cause a reversal of inhibition of fructose diphosphatase activity by AMP. The identification of phosphoenolpyruvate as an activator of a rate-limiting step in gluconeogenesis permits the suggestion of a coupling of the controlling mechanisms of several steps in the glycolytic and gluconeogenic chains. 7. These findings suggest mechanisms for the maintenance and regulation of control of fructose diphosphatase activity in king-crab skeletal muscle at low temperature and under conditions that favour concomitant activity of phosphofructokinase.

Project description:In this project, effects of chronic temperature changes on the transcriptome of the rainbow trout heart (Oncorhynchus mykiss) were examined. Ectothermic animals of north-temperate latitudes experience large seasonal changes in temperature which strongly affect the rate of body functions. To compensate for the effects of temperature changes ectotherms can respond to chronic temperature changes by increasing the quantity of tissue or enzyme needed for different physiological tasks, or by expressing proteins isoforms which are more appropriate for the new thermal conditions. On the other hand, proteins which are needed in lesser amounts in the new thermal regime could be depressed or down-regulated. Although expression of proteins can be changed by multiple mechanisms during synthesis and degradation, temperature dependent changes in transcription of genes is probably the most important factor in modifying the proteome of the tissues. Rainbow trout are active throughout their thermal tolerance range (0-25°C). Maintenance of adequate cardiac function at different thermal conditions requires a thorough change of the cardiac phenotype which appears as compensatory changes in relative heart mass, energy metabolism, nervous and humoral control of cardiac contractility and in electrical and mechanical properties of the trout heart. Such an extensive structural and functional remodeling of the heart probably necessitates both qualitative and quantitative changes among the numerous macromolecules which constitute the cardiac phenotype and cannot, therefore, be solely based on posttranslational modification of proteins but is expected to require differential gene expression. As a power supply of the circulatory system, the heart is in the focal point of physiological plasticity and sets limits for the activity level of the animal in different thermal conditions. Although several aspects of heart function have been studied in thermally acclimated fish and a number of structural changes have been noticed on exposure to different temperatures, the cellular and molecular mechanisms involved in chronic thermal stress of the fish heart are only partially elucidated. An interesting and poorly examined feature of the cold-acclimated phenotype of the trout heart is the increased heart mass which attenuates the depressive effect of low temperature on cardiac pump function. The heart is a complex organ composed of multiple tissues which together provide the system all necessary qualifications for cardiac pump function. In addition to the contractile and metabolic machinery of the cardiac myocytes, the amount and quality of the extracellular matrix also contribute to the properties of the heart as a muscular pump. Due to the complexity of the heart an enormous amounts of research efforts are needed to find out and examine all crucial aspects of cardiac plasticity required for thermal acclimation. In this regard the screening of gene expression by cDNA micro arrays might provide a broader view to genomic basis of cardiac remodeling under changing temperatures and aid to reveal the candidate genes which are important for thermal acclimation and which might remain unnoticed by traditional biochemical and physiological methods. In the present study we the steady-state effects of temperature acclimation on gene expression of the rainbow trout heart were screened by micro array analysis. Keywords: parallel sample

Project description:In this project, effects of chronic temperature changes on the transcriptome of the rainbow trout heart (Oncorhynchus mykiss) were examined. Ectothermic animals of north-temperate latitudes experience large seasonal changes in temperature which strongly affect the rate of body functions. To compensate for the effects of temperature changes ectotherms can respond to chronic temperature changes by increasing the quantity of tissue or enzyme needed for different physiological tasks, or by expressing proteins isoforms which are more appropriate for the new thermal conditions. On the other hand, proteins which are needed in lesser amounts in the new thermal regime could be depressed or down-regulated. Although expression of proteins can be changed by multiple mechanisms during synthesis and degradation, temperature dependent changes in transcription of genes is probably the most important factor in modifying the proteome of the tissues. Rainbow trout are active throughout their thermal tolerance range (0-25Â°C). Maintenance of adequate cardiac function at different thermal conditions requires a thorough change of the cardiac phenotype which appears as compensatory changes in relative heart mass, energy metabolism, nervous and humoral control of cardiac contractility and in electrical and mechanical properties of the trout heart. Such an extensive structural and functional remodeling of the heart probably necessitates both qualitative and quantitative changes among the numerous macromolecules which constitute the cardiac phenotype and cannot, therefore, be solely based on posttranslational modification of proteins but is expected to require differential gene expression. As a power supply of the circulatory system, the heart is in the focal point of physiological plasticity and sets limits for the activity level of the animal in different thermal conditions. Although several aspects of heart function have been studied in thermally acclimated fish and a number of structural changes have been noticed on exposure to different temperatures, the cellular and molecular mechanisms involved in chronic thermal stress of the fish heart are only partially elucidated. An interesting and poorly examined feature of the cold-acclimated phenotype of the trout heart is the increased heart mass which attenuates the depressive effect of low temperature on cardiac pump function. The heart is a complex organ composed of multiple tissues which together provide the system all necessary qualifications for cardiac pump function. In addition to the contractile and metabolic machinery of the cardiac myocytes, the amount and quality of the extracellular matrix also contribute to the properties of the heart as a muscular pump. Due to the complexity of the heart an enormous amounts of research efforts are needed to find out and examine all crucial aspects of cardiac plasticity required for thermal acclimation. In this regard the screening of gene expression by cDNA micro arrays might provide a broader view to genomic basis of cardiac remodeling under changing temperatures and aid to reveal the candidate genes which are important for thermal acclimation and which might remain unnoticed by traditional biochemical and physiological methods. In the present study we the steady-state effects of temperature acclimation on gene expression of the rainbow trout heart were screened by micro array analysis.

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Temperature and the regulation of enzyme activity in poikilotherms. Properties of rainbow-trout fructose diphosphatase.

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