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Pyruvate kinase variants of the Alaskan king-crab. Evidence for a temperature-dependent interconversion between two forms having distinct and adaptive kinetic properties.

ABSTRACT: 1. Pyruvate kinase of Alaskan king-crab leg muscle exists in two kinetically distinct forms, each of which displays a different temperature-dependence in the K(m) for phosphoenolpyruvate. 2. A ;cold' variant of the enzyme has hyperbolic kinetics and exhibits a minimal K(m) for substrate at 5 degrees . At physiological concentrations of phosphoenolpyruvate the ;cold' enzyme is active only below 10 degrees . A ;warm' pyruvate kinase has a minimal K(m) for substrate at about 12 degrees . This enzyme displays sigmoidal kinetics and is likely to be inactive, at physiological substrate concentrations, at temperatures below 9 degrees . 3. The combined activities of these two pyruvate kinases yield highly temperature-independent rates of catalysis, at physiological substrate concentrations, over the range of habitat temperatures encountered by the organism, namely 4-12 degrees . 4. The two variants of pyruvate kinase do not appear to be isoenzymes in the conventional sense. Electrophoretic and electrofocus analyses revealed only single peaks of activity. 5. The results suggest that the ;warm' pyruvate kinase and the ;cold' pyruvate kinase are formed by a temperature-dependent interconversion of one protein species. This interconversion has major adaptive significance: as the temperature is lowered the ;warm' enzyme is converted into the ;cold' enzyme; the opposite situation obtains when the temperature is raised. Temperature changes thus mimic the effects noted for fructose 1,6-diphosphate on certain mammalian pyruvate kinases.

SUBMITTER: Somero GN

PROVIDER: S-EPMC1184848 | biostudies-other | 1969 Sep

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.

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Pyruvate kinase variants of the Alaskan king-crab. Evidence for a temperature-dependent interconversion between two forms having distinct and adaptive kinetic properties.

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