| Résumé | Acetolactate synthase (ALS, AHAS; EC 4. 1. 3. 18) from Brassica napus has been partially purified and characterized using polyclonal antibodies. Following denaturing sodium dodecyl sulphate polyacrylamide gel electrophoresis and western blot analysis, 65 and 66 kDa ALS subunit polypeptides were immunologically detected, along with a novel 36 kDa polypeptide which cross-reacted with the anti-ALS antibody. Partial peptide sequencing of the 36 kDa peptide revealed significant similarity to plant aldolase proteins. ALS activity from stromal extracts fractionated by gel filtration chromatography as a single species of estimated molecular mass of 124 kDa, while comparative sedimentation coefficient in glycerol gradients indicated a corresponding molecular mass of 132 kDa. The results suggest that the native enzyme is a dimer of 65 and/or 66 kDa subunits. Anion exchange chromatography resolved two classes of ALS activity of equal native molecular weight, but which exhibited different properties with respect to subunit structure, sensitivity to inhibition by chlorsulfuron and feedback inhibition by branched chain amino acids. |
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