Effect of Casein on the Extractability and Stability of Nitrate Reductase From Wheat Leaves

The effect of casein on the extraction and stability of nitrate reductase (EC 1.6.6.1) from the primary leaf of the wheat (Triticum aestivum L.) cultivar Olympic was examined. The use of 2 per cent casein in the extraction medium resulted in a small but significant (P > 0·05) increase in nitrate reductase activity at the time of assay. This increase in activity was not attributable to improved extraction but was a result of increased stability of the enzyme between time of extraction and assay. The presence of casein actually resulted in less activity being extracted during the grinding procedure. Other cultivars of wheat examined showed a small increase in activity at the time of assay when extracted in the presence of casein. However, the ranking of these cultivars with respect to nitrate reductase activity did not alter markedly when casein was used. A number of other species examined showed a wide variation in response to casein, while buckwheat (Fagopyrum esculenta) and oats (Avena sativa) had an absolute requirement, maize (Zea mays cv. XL45) was relatively unaffected. Nitrate reductase in crude extracts decayed exponentially with time at both 0° and 10 °C with a half-life of 216 and 109 min respectively. Casein added before or after extraction completely stabilized the enzyme over a period of several hours at 0 °C while at 10 °C the enzyme had a half-life of approximately 16 h. The effect of casein at both 0° and 10 °C was independent of the concentration used within the range 0·5 to 4·0 per ant.