Partial purification and preliminary characterization of pyruvate kinase from the brown algaAscophyllum nodosum

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Abstract
The enzyme pyruvate kinase (EC 2.7.1.40) was partially purified from the marine brown alga Ascophyllum nodosum by means of DEAE-cellulose anion-exchange chromatography. The pH optimum for activity of this enzyme was 7·0 and the activation energy 62·2 kJ mol-1. In common with pyruvate kinase from other plants, the enzyme had a requirement for both K+ and Mg2+ (K a 2·46 and 1·15 mm, respectively). Substrate kinetics for ADP were hyperbolic (K m 0·15 mm), but those for phosphoenolpyruvate were sigmoid with a distinct positive cooperativity (K 0.5 1·68 mm, Hill coefficient 2·05). Fructose-6-phosphate (10 mm) acted as an allosteric activator, restoring Michaelis-Menten type kinetics for phosphoenolpyruvate (Hill coefficient 1·12) and substantially decreasing the K m (0·259 mm). Ascophyllum nodosum pyruvate kinase was also activated by AMP, and inhibited by ATP, CaCl2, citrate and oxalate.