Partial purification and properties of acetolactate synthase of etiolated pea seedlings

Two molecular species of ALS were found in etiolated pea seedlings on gel filtration column chromatography. Molecular weight of these two ALS were approximately 320,000 and 120,000 respectively. Higher molecular weight species designated large ALS was sensitive to the feedback inhibition by Leu/Val combination, while lower molecular species designated small ALS was insensitive. Large ALS which was fractionated by gel filtration column chromatography produced small ALS during the further chromatography of the same column. Km value for pyruvate and sensitivity to the feedback inhibition of large ALS were similar to that of the crude enzyme preparation. Both molecular species were sensitive to the inhibition by ALS-inhibiting herbicides. From these results, large ALS is considered to be the native enzyme in etiolated pea seedlings and to change to small ALS by loss of the regulatory center(s) during purification. This ALS showed the following enzymatical properties which had not been reported yet or were making a contrast to those in preceding papers. The rate of acetolactate production did not show a sigmoidal relationship with pyruvate concentration. Negative cooperativities were observed in the inhibition of ALS by leucine, valine and isoleucine. In contrast to Leu/Val combination and Leu/Ile combination, the antagonistic effect was found in the inhibition by Val/Ile combination. The inhibition type of Leu/Val combination varied depending on pyruvate concentration. The inhibitions by leucine or Leu/Val combination were desensitized by neither SH-inhibitors nor ATP