THE EFFECTS OF TEMPERATURE AND PH ON THE APPARENT MICHAELIS CONSTANT OF GLUTATHIONE-REDUCTASE FROM MAIZE (ZEA-MAYS L)

The interacting effects of temperature and pH on the kinetics of glutathione reductase from maize have been studied in detail. The apparent K(m) for oxidized glutathione (GSSG) measured with desalted crude extracts increased in an exponential manner with rising temperature as a single variable. Increasing pH as a single variable also resulted in higher values of apparent K(m) for GSSG. When pH was allowed to vary with temperature, a curve which combined the pH and temperature responses was observed. Temperature had the stronger influence and this combined curve was displaced from the temperature curve due to the effect of pH. The pH to which the assay buffer was adjusted at 30-degrees-C had an influence on the pattern of the results in this type of experiment. The response of apparent K(m) for NADPH, and of apparent K(m) for GSSG using partially-purified extracts, were also examined. The variation with temperature, at constant pH, was again exponential. The pattern of change of apparent K(m) with temperature is strongly dependent on experimental conditions. Affinity/temperature relationships deduced from such data would only reflect enzyme function in vivo if the physiological environment could be reproduced exactly in the assay mixture.